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WO2016187667A1 - Functionalised and substituted indoles as anti-cancer agents - Google Patents

Functionalised and substituted indoles as anti-cancer agents Download PDF

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Publication number
WO2016187667A1
WO2016187667A1 PCT/AU2016/050407 AU2016050407W WO2016187667A1 WO 2016187667 A1 WO2016187667 A1 WO 2016187667A1 AU 2016050407 W AU2016050407 W AU 2016050407W WO 2016187667 A1 WO2016187667 A1 WO 2016187667A1
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Prior art keywords
compound
dimethyl
methyl
indol
methylpiperazin
Prior art date
Application number
PCT/AU2016/050407
Other languages
French (fr)
Inventor
Eleanor Eiffe
Andrew Heaton
Peter Gunning
Herbert Treutlein
Jun Zeng
Ian James
Ian Dixon
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Novogen Limited
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Publication date
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Publication of WO2016187667A1 publication Critical patent/WO2016187667A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/496—Non-condensed piperazines containing further heterocyclic rings, e.g. rifampin, thiothixene or sparfloxacin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A61K31/403—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil condensed with carbocyclic rings, e.g. carbazole
    • A61K31/404—Indoles, e.g. pindolol
    • A61K31/4045—Indole-alkylamines; Amides thereof, e.g. serotonin, melatonin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/475—Quinolines; Isoquinolines having an indole ring, e.g. yohimbine, reserpine, strychnine, vinblastine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
    • C07D209/04—Indoles; Hydrogenated indoles
    • C07D209/08—Indoles; Hydrogenated indoles with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, directly attached to carbon atoms of the hetero ring
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the present invention relates broadly to functionalised and substituted indoles, the preparation thereof and their use in the treatment of proliferative diseases such as cancer.
  • patients with breast cancer have benefited from early screening programs as well as a variety of surgical techniques. However, these often prove physically and emotionally debilitating.
  • patients who have undergone surgery and subsequent chemotherapy often experience a recurrence in their disease.
  • a potential new method of specifically attacking cancer cells is through disruption of cancer cells' cellular skeletal system which is comprised predominantly of actin.
  • the actin cytoskeleton is intimately involved in cell division and cell migration.
  • actin plays a ubiquitous role as the cytoskeleton of tumor cells and the actin filaments of the muscle sarcomere.
  • the differing roles but similarity in structure make actin a difficult target for drug development due to unwanted off- target side effects.
  • the invention seeks to address one or more of the above mentioned problems, and/or to provide improvements in cancer therapy.
  • the present invention provides a compound of the formula (I),
  • R 1 and R 2" are independently selected from the group consisting of hydrogen and Ci- C 6 alkyl
  • R 3 J is NR 8°R 1 1 0 U or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR 6 and wherein the ring is optionally substituted with R ;
  • R 4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR 6 ;
  • each Y is independently OH or Ci-C 6 alkoxy and X is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 and wherein the ring is optionally substituted with between 1 and 3 substituents selected from the group consisting of: halo and Ci-C 6 alkyl, or
  • R 5 is wherein Z is OH or Ci-C 6 alkoxy and Q is selected from the group consisting of: halo, -S0 2 Ci-C 6 alkyl, -(CH 2 )o- 5 COOH, -(CH ⁇ o-sCOOQ-Ce alkyl or a 5- or 6-membered carbocyclic ring wherein between 1 and 4 carbons may optionally be replaced by N or NR 6 ;
  • R 6 is selected from the group consisting of: H and Ci-C 6 alkyl; R is selected from the group consisting of: H, halo, Ci-C 6 alkyl, Ci-C 6 alkoxy, CN, CF 3 and OCF 3 ;
  • R 8° and R 1 1 0 U are independently selected from the group consisting of: H and Ci-C 6 alkyl;
  • X 1 is an alkanediyl group having between 1 and 20 carbon atoms
  • X 3 are independently selected from the group consisting of: -0-, -NH-, -NCi- C 6 alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- or an alkanediyl group having between 1 and 20 carbon atoms wherein between 1 and 3 hydrogen atoms may optionally be replaced with R .
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of formula (I) according to the first aspect together with a pharmaceutically acceptable carrier, diluent or excipient.
  • the present invention provides a method for the treatment of a proliferative disease in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) according to the first aspect, or a pharmaceutical composition according to the second aspect.
  • the present invention provides use of a compound of formula (I) according to the first aspect in the manufacture of a medicament for the treatment of a proliferative disease.
  • the present invention provides use of a compound of formula (I) according to the first aspect or a pharmaceutical composition according to the second aspect for the treatment of a proliferative disease in a subject.
  • the present invention provides a compound of formula (I) according to the first aspect or a pharmaceutical composition according to the second aspect for use in the treatment of a proliferative disease in a subject.
  • the proliferative disease may be cancer, such as for example a solid tumour.
  • the cancer is selected from the group consisting of: breast cancer, lung cancer, prostate cancer, ovarian cancer, uterine cancer, brain cancer, skin cancer, colon cancer, bladder cancer, melanoma and neuroblastoma.
  • the cancer may be a cancer that has recurred.
  • the present invention provides a method for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence, the method comprising administration to the subject of an effective amount of a compound of formula (I) according to the first aspect, or a pharmaceutical composition according to the second aspect.
  • the present invention provides use of a compound of formula (I) according to the first aspect in the manufacture of a medicament for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
  • the present invention provides a compound of formula (I) according to the first aspect for use in reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
  • the present invention provides use of a compound of formula (I) according to the first aspect for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
  • the subject may be a subject who is in cancer remission.
  • the present invention provides a combination comprising:
  • the combination may be a synergistic combination.
  • the present invention provides a pharmaceutical composition comprising:
  • composition (iii) a pharmaceutically acceptable carrier, diluent or excipient.
  • the composition may be a synergistic composition.
  • the present invention provides a kit comprising:
  • compositions are intended for simultaneous, concurrent, separate or sequential use.
  • the present invention provides a method for the treatment of cancer in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) according to the first aspect, and a therapeutically effective amount of vincristine or paclitaxel.
  • the cancer may be melanoma, lung cancer or prostate cancer.
  • Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential.
  • the compound of formula (I) may be selected from one or more of compounds 6, 8, 14, 16, 23, 24, 27, 31, 32, 38, 39, 47, 48, 51, 55, 58, 60, 63, 67 and 68.
  • an element means one element or more than one element.
  • alkyl is taken to mean straight chain or branched chain monovalent saturated hydrocarbon groups having the recited number of carbon atoms.
  • alkyl groups include, but are not limited to, methyl, ethyl, 1 -propyl, isopropyl, 1- butyl, 2-butyl, isobutyl, tert-butyl, amyl, 1,2-dimethylpropyl, 1,1-dimethylpropyl, pentyl, isopentyl, hexyl, 4-methylpentyl, 1-methylpentyl, 2-methylpentyl, 3- methylpentyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl, 1,2-dimethylbutyl, 1,3- dimethylbutyl, 1,2,2-trimethylpropyl, 1,1,2-trimethylpropyl, 2-ethylpentyl, 3- ethylpentyl, heptyl, 1-methylhexyl, 2,
  • alkoxy is taken to mean O-alkyl groups in which alkyl is as defined herein.
  • alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, sec-butoxy and tert-butoxy.
  • halogen and halo are synonymous and refer to fluorine, chlorine, bromine or iodine.
  • alkanediyl is understood to refer to a bivalent saturated branched or straight chain hydrocarbon group conforming to the formula C n H 2n .
  • carrier ring refers to a ring structure in which all of the ring members are carbon atoms.
  • pharmaceutically acceptable salt refers to those salts which, within the scope of sound medical judgement, are suitable for use in contact with the tissues of humans and animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • prodrug means a compound which is able to be converted in vivo by metabolic means (e.g. by hydrolysis, reduction or oxidation) to a compound of the formula (I).
  • metabolic means e.g. by hydrolysis, reduction or oxidation
  • an ester prodrug of a compound of the present invention containing a hydroxy group may be hydrolysed in vivo to the parent molecule.
  • Suitable esters are for example, acetates, citrates, lactates, tartrates, malonates, oxalates, salicylates, propionates, succinates, fumarates, maleates, methylene-bis- ⁇ - hydroxynaphthoates, gestisates, isethionates, di-/?-toluoyltartrates, methanesulfonates, ethanesulfonates, benzenesulfonates, /?-toluenesulfonates, cyclohexylsulfamates and quinates.
  • treating refers to any and all uses which remedy a proliferative disease or symptoms thereof, prevent the establishment of a proliferative disease, or otherwise prevent, hinder, retard or reverse the progression of a proliferative disease or other undesirable symptoms in any way whatsoever.
  • treating does not necessarily imply that a subject is treated until total recovery.
  • terapéuticaally effective amount includes a non-toxic but sufficient amount of an active compound to provide the desired therapeutic effect. Those skilled in the art will appreciate that the exact amount of a compound required will vary based on a number of factors and thus it is not possible to specify an exact “therapeutically effective amount”. However, for any given case an appropriate “therapeutically effective amount” may be determined by one of ordinary skill in the art.
  • an effective amount includes a non-toxic but sufficient amount of an active compound to provide the stated effect.
  • effective amount means an amount of a compound of formula (I) that is required to reduce the incidence of, or risk of an individual experiencing cancer recurrence.
  • Those skilled in the art will appreciate that the exact amount of a compound required will vary based on a number of factors and thus it is not possible to specify an exact "effective amount”. However, for any given case an appropriate “effective amount” may be determined by one of ordinary skill in the art.
  • a “pharmaceutical carrier, diluent or excipient” includes, but is not limited to, any physiological buffered (i.e., about pH 7.0 to 7.4) medium comprising a suitable water-soluble organic carrier, conventional solvents, dispersion media, fillers, solid carriers, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents.
  • suitable water-soluble organic carriers include, but are not limited to saline, dextrose, corn oil, dimethylsulfoxide, and gelatin capsules.
  • lactose lactose
  • mannitol corn starch
  • potato starch binders such as crystalline cellulose, cellulose derivatives, acacia, gelatins, disintegrators, such as sodium carboxymethyl-cellulose, and lubricants such as talc or magnesium stearate.
  • binders such as crystalline cellulose, cellulose derivatives, acacia, gelatins
  • disintegrators such as sodium carboxymethyl-cellulose
  • lubricants such as talc or magnesium stearate.
  • subject includes any human or non-human animal.
  • the compounds of the present invention may also be useful for veterinary treatment of mammals, including companion animals and farm animals, such as, but not limited to dogs, cats, horses, cows, sheep, and pigs.
  • the term "recurrence" as it relates to cancer is understood to mean the return of cancerous cells and/or a cancerous tumour after cancerous cells and/or a cancerous tumour have been successfully treated previously.
  • the term “administering” and variations of that term including “administer” and “administration”, include contacting, applying, delivering or providing a compound or composition of the invention to an organism, or a surface by any appropriate means.
  • Figure 1 Imaging and quantitation of actin filaments in SK-N-SH neuroblastoma cells treated with compound (A) 50, (B) 51.
  • Cells were stained with 488-Atto-Phallodin and DAPI to visualize the actin filament bundles and the nucleus, respectively.
  • Shown in the top panel is a representative gray scale immunofluorescent image from control (vehicle alone), 2.5 ⁇ and 5 ⁇ treated cells, overlayed with the linear feature quantitation. The coloured lines indicate the detected actin filaments.
  • Also shown is the quantitation of cell number and filament number/cell.
  • Statistical analysis was performed using a one way ANNOVA-multiple comparison where each drug-treated group was compared to the control. **** p ⁇ 0.0001, **** p ⁇ 0.001, *** p ⁇ 0.01, ** p ⁇ 0.1.
  • FIG. 2 Imaging and quantitation of actin filaments in SK-N-SH neuroblastoma cells treated with compound (A) 50 and (B) 51.
  • Cells were stained with j9d primary antibody (MAb culture s/n clone 2G10.2, 1 :50) followed by 488- conjugated secondary antibody (Goat anti mouse 488, 1: 1000) and DAPI to visualize the Tpm3.1 containing filament bundles and the nucleus, respectively.
  • Shown in the top panel (enlarged inset bottom panel) is a representative gray scale immunofluorescent image from control (vehicle alone), 2.5 ⁇ and 5 ⁇ treated cells, overlayed with the linear feature quantitation. The coloured lines indicate the detected actin filaments.
  • FIG. 3 Plate layout for drug combination assays. Each screening plate contained up to six 6x6 dose matrix along with assay controls (thonzonium bromide and DMSO) as indicated. The rest of the wells contained cells only. Individual drug doses within the matrix were based on the established IC 50 concentrations for the two drugs using two-fold dilution steps. Each drug combination was tested in triplicate.
  • the present invention is based on the surprising finding by the inventors that compounds of the general formula (I) effectively inhibit tropomyosin which results in an unexpected improvement in the treatment of proliferative diseases, particularly cancer.
  • actin cytoskeleton involves a number of ancillary control and regulatory proteins. Identification and specific targeting of actin regulatory proteins associated with the cytoskeleton of cancer cells provides the opportunity to develop cancer- specific drugs lacking unwanted side effects.
  • Actin filaments are constructed through the polymerisation of globular actin protein monomers.
  • the actin monomer is polar, with one end bearing a positive charge and the other end a negative charge.
  • the actin filaments thus have all the actin proteins aligned in one direction.
  • These filaments have secondary coiled proteins, tropomyosins, associated with them.
  • the tropomyosins play an integral role in regulating the function of actin filaments.
  • the actin filaments are made up of polymeric actin monomers with tropomyosin dimers sitting in the alpha helical groove of the actin filament to form a homopolymer.
  • tropomyosin isoforms there are more than 40 mammalian tropomyosin isoforms, each of which regulates specific actin filaments.
  • tropomyosins that regulate the cytoskeleton of cancer cells; disruption of this interaction offers a basis to treat cancer cells with a high degree of specificity.
  • the present invention provides a compound of the formula (I),
  • R 1 and R 2" are independently selected from the group consisting of hydrogen and Ci- C 6 alkyl
  • R 3 J is NR 8°R 1 1 0 U or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR 6 and wherein the ring is optionally substituted with R ;
  • R 4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR 6 ;
  • each Y is independently OH or Ci-C 6 alkoxy and X is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 and wherein the ring is optionally substituted with between 1 and 3 substituents selected from the group consisting of: halo and Ci-C 6 alkyl, or
  • R is Q wherein Z is OH or Ci-C 6 alkoxy and Q is selected from the group consisting of: halo, -S0 2 Ci-C 6 alkyl, -(CH 2 )o- 5 COOH, -(CH ⁇ o-sCOOQ-Ce alkyl or a 5- or 6-membered carbocyclic ring wherein between 1 and 4 carbons may optionally be replaced by N or NR 6 ;
  • R 6 is selected from the group consisting of: H and Ci-C 6 alkyl; R is selected from the group consisting of: H, halo, Ci-C 6 alkyl, Ci-C 6 alkoxy, CN, CF 3 and OCF 3 ;
  • R 8° and R 1 1 0 U are independently selected from the group consisting of: H and Ci-C 6 alkyl;
  • X 1 is an alkanediyl group having between 1 and 20 carbon atoms
  • X 2" and X 3 3 are independently selected from the group consisting of: -0-, -NH-, -NCi- C 6 alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- or an alkanediyl group having between 1 and 20 carbon atoms wherein between 1 and 3 hydrogen atoms may optionally be replaced with R .
  • R 1 and R 2 are independently selected from the group consisting of: H and Ci-C 3 alkyl.
  • R 1 and R 2 are independently selected from the group consisting of: H and methyl.
  • R 1 and R 2 are both methyl.
  • R 3 is NR 8 R 10 or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 and wherein the ring is optionally substituted with R .
  • R 3 is NR 8 R 10 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 and wherein the ring is optionally substituted with Ci-C 6 alkyl.
  • R 3 is NR 8 R 10 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 .
  • R is N(Me) 2 , NH 2 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR 6 .
  • R is N(Me) 2 , NH 2 or a 5- or 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N or NR 6 .
  • R is N(Me) 2 , NH 2 or a 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
  • R is N(Me) 2 or a 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
  • R is N(Me) 2 , NH 2 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, S, O or NR 6 .
  • the 4- to 7-membered carbocyclic ring and the 5- or 6- membered carbocyclic ring may be saturated or partially saturated.
  • R is selected from the group consisting of: N(Me) 2 ,
  • R is selected from the group consisting of: N(Me) 2
  • odiment R is selected from the group consisting of:
  • R 4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, NR 6 , O or S.
  • R 4 is phenylene or a 5- or 6-membered heteroarylene ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S.
  • R 4 is phenylene or a 5- or 6-membered heteroarylene ring having between 1 and 3 N heteroatoms.
  • R 4 is phenylene or a 6-membered heteroarylene ring having between 1 and 3 N heteroatoms.
  • R 4 is phenylene or pyridylene.
  • R 4 is phenylene
  • R 4 is para-phenylene or meta-phenylene.
  • R 5 is wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S,and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 substituents selected from the group consisting of: halo, methyl, ethyl, propyl and isopropyl, or
  • R is Q wherein Z is OH or OMe and Q is selected from the group consisting of: halo, -S0 2 Me, -(CH 2 ) 0 - 5 COOH, -(CH 2 ) 0 - 5 COOCi-C 6 alkyl, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
  • R 5 is Y is OH or OMe and X is or a 5- or 6- membered heteroaryl ring having between 1 and 3 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 halo substituents, or
  • R 5 is wherein Z is OH or OMe and Q is selected from the group consisting of: halo, -S0 2 Me, -(CH 2 )i- 5 COOH, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
  • R 5 is Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a halo substituent, or
  • R 5 is wherein Z is OH or OMe and Q is selected from the group consisting of: I, -S0 2 Me, -(CH 2 )i_ 3 COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms.
  • R 5 is wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or
  • R is Q wherein Z is OH and Q is selected from the group consisting of: I, -S0 2 Me, -(CH 2 ) 2 COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms.
  • R is Y is OH or OMe and
  • X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or
  • R is Q wherein Z is OH and Q is selected from the group consisting of: I, -S0 2 Me, -(CH 2 ) 2 COOH and tetrazolyl.
  • R 5 is
  • R 5 is
  • R 6 is selected from the group consisting of H, methyl and ethyl.
  • R 6 is selected from the group consisting of H and methyl.
  • X 1 is an alkanediyl group having between 1 and 15 carbon atoms.
  • X 1 is an alkanediyl group having between 1 and 10 carbon atoms.
  • X 1 is an alkanediyl group having between 1 and 6 carbon atoms.
  • X 1 is - (CH 2 ) 1-6 - or -CH 2 -CH(Me)-(CH 2 )-. In yet another embodiment X 1 is - (CH 2 ) 2 _ 5 - or -CH 2 -CH(Me)-(CH 2 )-.
  • X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NCi-C 6 alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 15 carbon atoms.
  • X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 10 carbon atoms.
  • X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 6 carbon atoms.
  • X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -C(0)NH-, -NHC(O)-, -CH 2 CH 2 CH 2 -, CH 2 CH 2 - and - CH 2 -.
  • X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -C(0)NH-, -NHC(O)- and CH 2 .
  • X is selected from the group consisting of: -0-, -NH- and -CH 2 -.
  • X 3 is -C(0)NH- or -NHC(O)-.
  • exemplary compounds of the formula (I) include the following:
  • the compounds are:
  • the compound of formula (I) is selected from the group consisting of: compounds 9-12, 17-20, 22-24, 30-32, 44, 46-52 and 69-72.
  • the compound of formula (I) is selected from the group consisting of: compounds 1-12, 14-20, 26-28, 33-36, 38-40, 42-44 and 46-72.
  • the compound of formula (I) is selected from the group consisting of: compounds 22-24 and 30-32. In yet another embodiment the compound of formula (I) is selected from the group consisting of: compounds 6, 8, 16, 23, 24, 47, 48, 51, 60 and 68.
  • the compounds of formula (I) may include one or more chiral centres.
  • the present invention includes all enantiomers and diastereoisomers as well as mixtures thereof in any proportions.
  • the invention also extends to isolated enantiomers or pairs of enantiomers. Methods of separating enantiomers and diastereoisomers are well known to persons skilled in the art.
  • compounds of the formula (I) are racemic mixtures.
  • compounds of the formula (I) are present in optically pure form.
  • Solvates are complexes formed by association of molecules of a solvent with a compound of the formula (I).
  • compounds of the formula (I) that are solids, it will be understood by those skilled in the art that such compounds may exist in different crystalline or polymorphic forms, all of which are intended to be within the scope of the present invention.
  • the compounds of formula (I) may be in the form of pharmaceutically acceptable salts. Such salts are well known to those skilled in the art. S. M. Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66: 1-19. Pharmaceutically acceptable salts can be prepared in situ during the final isolation and purification of compounds of the formula (I), or separately by reacting the free base compound with a suitable organic acid. Suitable pharmaceutically acceptable acid addition salts of the compounds of the present invention may be prepared from an inorganic acid or from an organic acid. Examples of such inorganic acids are hydrochloric, hydrobromic, hydroiodic, nitric, carbonic, sulfuric, and phosphoric acid.
  • Appropriate organic acids may be selected from aliphatic, cycloaliphatic, aromatic, heterocyclic carboxylic and sulfonic classes of organic acids, examples of which are formic, acetic, propionic, succinic, glycolic, gluconic, lactic, malic, tartaric, citric, ascorbic, glucoronic, fumaric, maleic, pyruvic, alkyl sulfonic, arylsulfonic, aspartic, glutamic, benzoic, anthranilic, mesylic, salicylic, p-hydroxybenzoic, phenylacetic, mandelic, ambonic, pamoic, pantothenic, sulfanilic, cyclohexylaminosulfonic, stearic, algenic, ⁇ -hydroxybutyric, galactaric, and galacturonic acids.
  • Suitable pharmaceutically acceptable base addition salts of the compounds of the present invention include metallic salts made from lithium, sodium, potassium, magnesium, calcium, aluminium, and zinc, and organic salts made from organic bases such as choline, diethanolamine, morpholine.
  • the compounds of formula (I) also extend to include all derivatives with physiologically cleavable leaving groups that can be cleaved in vivo to provide the compounds of the formula (I).
  • Suitable leaving groups include acyl, phosphate, sulfate, sulfonate, and preferably are mono-, di- and per-acyl oxy-substituted compounds, where one or more of the pendant hydroxy groups are protected by an acyl group, preferably an acetyl group.
  • acyloxy substituted compounds are readily cleavable to the corresponding hydroxy-substituted compounds.
  • compound (III) could be hydrolysed to give the free acid and subsequently reacted with R 5 NH 2 prior to functionalisation of the 1 -position of the indole to introduce the X 1 R 3 substituent.
  • This alternative synthetic route is shown in Scheme 2 below.
  • Compounds of the formula (I) may be used in the treatment or prevention of proliferative diseases such as cancer.
  • the compounds and pharmaceutical compositions of the invention may be useful for the treatment of a wide variety of cancers, including but not limited to, solid tumours such as breast cancer, lung cancer (NSCLC and SCLC), prostate cancer, ovarian cancer, uterine cancer, peritoneal cancer, brain cancer (including, for example, gliomas such as glioblastoma, Diffuse Intrinsic Pontine Glioma (DIPG) and medulloblastoma), skin cancer, colon cancer, bladder cancer, colorectal cancer, gastric cancer, liver cancer, pancreatic cancer, head and neck cancer, melanoma, malignant ascites, mesothelioma and neuroblastoma.
  • the cancer is selected from the group consisting of: prostate cancer, lung cancer, breast cancer, colorectal cancer, melanoma and neuroblastoma.
  • the cancer may be
  • Compounds of the formula (I) may also find use in treating cancer that has recurred and in reducing the incidence of, or the risk of, recurrence of cancer in a subject deemed to be at risk of cancer recurrence, for example a subject who is in cancer remission.
  • the subject may be in remission from a solid tumour as defined herein.
  • compounds of the formula (I) may possess superior pharmaceutical properties, such as improved resistance to conjugation via glucuronyl transferases and other water-solubilising transferases such as sulfases, which may be over-expressed on proliferative cells, such as cancer cells.
  • superior pharmaceutical properties such as an enhanced pharmacokinetic profile through reduced conjugation and elimination.
  • compounds and pharmaceutical compositions of the present invention may be administered via any route which delivers an effective amount of the compounds to the tissue or site to be treated.
  • the compounds and compositions may be administered by the parenteral (for example intravenous, intraspinal, subcutaneous or intramuscular), oral or topical route. Administration may be systemic, regional or local. In one embodiment administration may be rectal.
  • the particular route of administration to be used in any given circumstance will depend on a number of factors, including the nature of the cancer to be treated, the severity and extent of the cancer, the required dosage of the particular compound to be delivered and the potential side-effects of the compound.
  • suitable compositions may be prepared according to methods that are known to those of ordinary skill in the art and may include pharmaceutically acceptable carriers, diluents and/or excipients.
  • the carriers, diluents and excipients must be "acceptable” in terms of being compatible with the other ingredients of the composition, and not deleterious to the recipient thereof.
  • Examples of pharmaceutically acceptable carriers or diluents are demineralised or distilled water; saline solution; vegetable based oils such as peanut oil, safflower oil, olive oil, cottonseed oil, maize oil or coconut oil; silicone oils, including polysiloxanes, such as methyl polysiloxane, phenyl polysiloxane and methylphenyl polysiloxane; volatile silicones; mineral oils such as liquid paraffin, soft paraffin or squalane; cellulose derivatives such as methyl cellulose, ethyl cellulose, carboxymethylcellulose, sodium carboxymethylcellulose or hydroxypropylmethylcellulose; Cremaphor; cyclodextrins; lower alkanols, for example ethanol or i-propanol; lower aralkanols; lower polyalkylene glycols or lower alkylene glycols, for example polyethylene glycol, polypropylene glycol, ethylene glycol, propylene glycol, 1,3-butylene glyco
  • compositions of the invention may be in a form suitable for administration by injection, in the form of a formulation suitable for oral ingestion (such as capsules, tablets, caplets, elixirs, for example), in the form of an ointment, cream or lotion suitable for topical administration, in a form suitable for delivery as an eye drop, in an aerosol form suitable for administration by inhalation, such as by intranasal inhalation or oral inhalation, in a form suitable for parenteral administration, that is, subcutaneous, intramuscular or intravenous injection.
  • a formulation suitable for oral ingestion such as capsules, tablets, caplets, elixirs, for example
  • an ointment cream or lotion suitable for topical administration
  • an eye drop in an aerosol form suitable for administration by inhalation, such as by intranasal inhalation or oral inhalation
  • parenteral administration that is, subcutaneous, intramuscular or intravenous injection.
  • non-toxic parenterally acceptable diluents or carriers can include cyclodextrins (for example Captisol®) Cremaphor, Ringer's solution, isotonic saline, phosphate buffered saline, ethanol and 1,2 propylene glycol.
  • cyclodextrins for example Captisol®
  • the compounds may also be added to PEG and non-PEGylated liposomes or micelles with specific targeting tags attached to PEG moieties, such as the RGD peptide or glutathione, for aiding passage across the blood brain barrier.
  • suitable carriers, diluents, excipients and adjuvants for oral use include cyclodextrins, Cremaphor, peanut oil, liquid paraffin, sodium carboxymethylcellulose, methylcellulose, sodium alginate, gum acacia, gum tragacanth, dextrose, sucrose, sorbitol, mannitol, gelatine and lecithin.
  • these oral formulations may contain suitable flavouring and colourings agents.
  • the capsules When used in capsule form the capsules may be coated with compounds such as glyceryl monostearate or glyceryl distearate that delay disintegration.
  • Adjuvants typically include emollients, emulsifiers, thickening agents, preservatives, bactericides and buffering agents.
  • Solid forms for oral administration may contain binders acceptable in human and veterinary pharmaceutical practice, sweeteners, disintegrating agents, diluents, flavourings, coating agents, preservatives, lubricants and/or time delay agents.
  • Suitable binders include gum acacia, gelatine, corn starch, gum tragacanth, sodium alginate, carboxymethylcellulose or polyethylene glycol.
  • Suitable sweeteners include sucrose, lactose, glucose, aspartame or saccharin.
  • Suitable disintegrating agents include corn starch, methylcellulose, polyvinylpyrrolidone, guar gum, xanthan gum, bentonite, alginic acid or agar.
  • Suitable diluents include lactose, sorbitol, mannitol, dextrose, kaolin, cellulose, calcium carbonate, calcium silicate or dicalcium phosphate.
  • Suitable flavouring agents include peppermint oil, oil of wintergreen, cherry, orange or raspberry flavouring.
  • Suitable coating agents include polymers or copolymers of acrylic acid and/or methacrylic acid and/or their esters, waxes, fatty alcohols, zein, shellac or gluten.
  • Suitable preservatives include sodium benzoate, vitamin E, alpha-tocopherol, ascorbic acid, methyl paraben, propyl paraben or sodium bisulphite.
  • Suitable lubricants include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc.
  • Suitable time delay agents include glyceryl monostearate or glyceryl distearate.
  • Liquid forms for oral administration may contain, in addition to the above agents, a liquid carrier.
  • suitable liquid carriers include water, oils such as olive oil, peanut oil, sesame oil, sunflower oil, safflower oil, coconut oil, liquid paraffin, ethylene glycol, propylene glycol, polyethylene glycol, ethanol, propanol, isopropanol, glycerol, fatty alcohols, triglycerides or mixtures thereof.
  • Suspensions for oral administration may further comprise dispersing agents and/or suspending agents.
  • Suitable suspending agents include sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, sodium alginate or acetyl alcohol.
  • Suitable dispersing agents include lecithin, polyoxyethylene esters of fatty acids such as stearic acid, polyoxyethylene sorbitol mono- or di-oleate, -stearate or -laurate, polyoxyethylene sorbitan mono- or di-oleate, -stearate or -laurate and the like.
  • Emulsions for oral administration may further comprise one or more emulsifying agents.
  • Suitable emulsifying agents include dispersing agents as exemplified above or natural gums such as guar gum, gum acacia or gum tragacanth.
  • parenterally administrable compositions are apparent to those skilled in the art, and are described in more detail in, for example, Remington's Pharmaceutical Science, 15th ed., Mack Publishing Company, Easton, Pa., hereby incorporated by reference herein.
  • Topical formulations may comprise an active ingredient together with one or more acceptable carriers, and optionally any other therapeutic ingredients.
  • Formulations suitable for topical administration include liquid or semi-liquid preparations suitable for penetration through the skin to the site where treatment is required, such as liniments, lotions, creams, ointments or pastes, and drops suitable for administration to the eye, ear or nose.
  • Drops according to the present invention may comprise sterile aqueous or oily solutions or suspensions. These may be prepared by dissolving the active ingredient in an aqueous solution of a bactericidal and/or fungicidal agent and/or any other suitable preservative, and optionally including a surface active agent. The resulting solution may then be clarified by filtration, transferred to a suitable container and sterilised. Sterilisation may be achieved by autoclaving or maintaining at 90 °C to 100 °C for half an hour, or by filtration, followed by transfer to a container by an aseptic technique.
  • bactericidal and fungicidal agents suitable for inclusion in the drops are phenylmercuric nitrate or acetate (0.002%), benzalkonium chloride (0.01%) and chlorhexidine acetate (0.01%).
  • Suitable solvents for the preparation of an oily solution include glycerol, diluted alcohol and propylene glycol.
  • Lotions according to the present invention include those suitable for application to the skin or eye.
  • An eye lotion may comprise a sterile aqueous solution optionally containing a bactericide and may be prepared by methods similar to those described above in relation to the preparation of drops.
  • Lotions or liniments for application to the skin may also include an agent to hasten drying and to cool the skin, such as an alcohol or acetone, and/or a moisiteriser such as glycerol, or oil such as olive oil.
  • Creams, ointments or pastes according to the present invention are semi-solid formulations of the active ingredient for external application. They may be made by mixing the active ingredient in finely divided or powdered form, alone or in solution or suspension in an aqueous or non-aqueous fluid, with a greasy or non-greasy basis.
  • the basis may comprise hydrocarbons such as hard, soft or liquid paraffin, glycerol, beeswax, a metallic soap; a mucilage; an oil of natural origin such as almond, corn, arachis, castor or olive oil; wool fat or its derivatives, or a fatty acid such as stearic or oleic acid together with an alcohol, such as propylene glycol or macrogols.
  • composition may incorporate any suitable surfactant such as an anionic, cationic or non-ionic surfactant, such as sorbitan esters or polyoxyethylene derivatives thereof.
  • suitable surfactant such as an anionic, cationic or non-ionic surfactant, such as sorbitan esters or polyoxyethylene derivatives thereof.
  • Suspending agents such as natural gums, cellulose derivatives or inoraganic materials such as silicaceous silicas, and other ingredients such a lanolin, may also be included.
  • compositions are administered in the form of suppositories suitable for rectal administration of the compounds of formula (I).
  • suppositories suitable for rectal administration of the compounds of formula (I).
  • These compositions are prepared by mixing the compound of formula (I) with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the compound of formula (I).
  • suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the compound of formula (I).
  • suitable non-irritating excipient include cocoa butter, glycerinated gelatin, hydrogenated vegetable oils, mixtures of polyethylene glycols of various molecular weights and fatty acid esters of polyethylene glycol.
  • compositions may also be administered or delivered to target cells in the form of liposomes.
  • Liposomes are generally derived from phospholipids or other lipid substances and are formed by mono- or multi-lamellar hydrated liquid crystals that are dispersed in an aqueous medium.
  • liposomes used in administering or delivering a composition to target cells are synthetic cholesterol (Sigma), the phospholipid l,2-distearoyl-sn-glycero-3-phosphocholine (DSPC); Avanti Polar Lipids), the PEG lipid 3-N-[(-methoxy poly(ethylene glycol)2000)carbamoyl]-l,2-dimyrestyloxy-propylamine (PEG-cDMA), and the cationic lipid l,2-di-o-octadecenyl-3-(N,N-dimethyl)aminopropane (DODMA) or 1,2- dilinoleyloxy-3-(N,N-dimethyl)aminopropane (DLinDMA) in the molar ratios 55:20: 10: 15 or 48:20:2:30, respectively, PEG-cDMA, DODMA and DLinDMA.
  • DSPC phospholipid l,2-distearoyl
  • the liposome may be contracted from l,2-distearoyl-sn-glycero-3-phosphoethanolamine- N-[methoxy(polyethylene glycol)-2000] (DSPE PEG2000) and phosphatidylcholine derived from soy and hydrogenated between 50-100%, for example Soy PC-75 or Soy PC- 100. Differing MW PEG's may be used and covalently bound with various specific targeting agents such as glutathione, RGD peptides or other recognized liposome targeting agents. Any non-toxic, physiologically acceptable and metabolisable lipid capable of forming liposomes can be used.
  • the compositions in liposome form may contain stablisers, preservatives, excipients and the like.
  • the preferred lipids are the phospholipids and the phosphatidyl cholines (lecithins), both natural and synthetic.
  • Methods to form liposomes are known in the art, and in relation to this, specific reference is made to: Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, N.Y. (1976), p. 33 et seq., the contents of which is incorporated herein by reference.
  • compositions may also be administered in the form of microparticles or nanoparticles.
  • Biodegradable microparticles formed from polyactide (PLA), polylactide-co-glycolide (PLGA), and epsilon-caprolactone ( ⁇ -caprlactone) have been extensively used as drug carriers to increase plasma half life and thereby prolong efficacy (R. Kumar, M., 2000, J. Pharm. Pharmaceut. Sci. 3(2) 234-258).
  • Microparticles have been formulated for the delivery of a range of drug candidates including vaccines, antibiotics, and DNA. Moreover, these formulations have been developed for various delivery routes including parenteral subcutaneous injection, intravenous injection and inhalation.
  • compositions may incorporate a controlled release matrix that is composed of sucrose acetate isobutyrate (SAIB) and an organic solvent or organic solvents mixture.
  • SAIB sucrose acetate isobutyrate
  • Polymer additives may be added to the vehicle as a release modifier to further increase the viscosity and slow down the release rate.
  • SAIB is a well known food additive. It is a very hydrophobic, fully esterified sucrose derivative, at a nominal ratio of six isobutyrate to two acetate groups. As a mixed ester, SAIB does not crystallise but exists as a clear viscous liquid. Mixing SAIB with a pharmaceutically acceptable organic solvent, such as ethanol or benzyl alcohol decreases the viscosity of the mixture sufficiently to allow for injection.
  • An active pharmaceutical ingredient may be added to the SAIB delivery vehicle to form SAIB solution or suspension formulations.
  • the solvent differs from the matrix allowing the SAIB-drug or SAIB- drug-polymer mixtures to set up as an in situ forming depot.
  • compositions may be administered to subjects either therapeutically or preventively.
  • compositions are administered to a patient already suffering from cancer in an amount sufficient to cure or at least partially arrest the cancer and its complications.
  • the composition should provide a quantity of the compound or agent sufficient to effectively treat the subject.
  • the therapeutically effective amount for any particular subject will depend upon a variety of factors including: the cancer being treated and the severity thereof; the activity of the compound administered; the composition in which the compound is present; the age, body weight, general health, sex and diet of the subject; the time of administration; the route of administration; the rate of sequestration of the compound; the duration of the treatment; drugs used in combination or coincidental with the compound, together with other related factors well known in medicine.
  • an effective dosage is expected to be in the range of about 0.0001 mg to about 1000 mg per kg body weight per 24 hours; typically, about 0.001 mg to about 750 mg per kg body weight per 24 hours; about 0.01 mg to about 500 mg per kg body weight per 24 hours; about 0.1 mg to about 500 mg per kg body weight per 24 hours; about 0.1 mg to about 250 mg per kg body weight per 24 hours; about 1.0 mg to about 250 mg per kg body weight per 24 hours.
  • an effective dose range is expected to be in the range about 1.0 mg to about 200 mg per kg body weight per 24 hours; about 1.0 mg to about 100 mg per kg body weight per 24 hours; about 1.0 mg to about 50 mg per kg body weight per 24 hours; about 1.0 mg to about 25 mg per kg body weight per 24 hours; about 5.0 mg to about 50 mg per kg body weight per 24 hours; about 5.0 mg to about 20 mg per kg body weight per 24 hours; about 5.0 mg to about 15 mg per kg body weight per 24 hours.
  • an effective dosage may be up to about 500 mg/m .
  • an effective dosage is expected to be in the range of about 25 to about 500 mg/m , preferably about 25 to about 350 mg/m , more preferably about 25 to about 300
  • the treatment would be for the duration of the disease state.
  • the compounds of formula (I) may be used alone in the treatment of cancer, or alternatively in combination with radiotherapy and/or surgery and/or other therapeutic agents, for example chemotherapeutic agents and immunostimulatory agents, as part of a combination therapy.
  • the compounds of formula (I) may sensitise cancer cells to other chemotherapeutic agents and/or radiotherapy.
  • combination therapy and "adjunct therapy” are intended to embrace administration of multiple therapeutic agents in a sequential manner in a regimen that will provide beneficial effects and is intended to embrace administration of these agents in either a single formulation or in separate formulations.
  • Combination therapy may involve the active agents being administered together, sequentially, or spaced apart as appropriate in each case.
  • Combinations of active agents including compounds of the invention may be synergistic.
  • co-administration of compounds of the formula (I) with other therapeutic agent(s) may be effected by a compound of the formula (I) being in the same unit dose form as the other therapeutic agent(s), or the compound of the formula (I) and the other therapeutic agent(s) may be present in individual and discrete unit dosage forms that are administered concurrently, sequentially or separately.
  • Separate administration refers to administration of the compound of the formula (I) independently of the other therapeutic agent(s) at different time points. It may be the case that the compound of the formula (I) and the other therapeutic agent(s) are administered such that no overlap of measurable blood levels of the compound formula (I) and the other therapeutic agent(s) are present.
  • sequence administration may be in any order as required, and may require an ongoing physiological effect of the first or initial agent to be current when the second or later agent is administered, especially where a cumulative or synergistic effect is desired.
  • one or more compounds of formula (I) may be included in combination therapy with surgery and/or radiotherapy and/or one or more chemotherapeutic agents.
  • chemotherapeutic agents that are currently in use, in clinical evaluation and in pre-clinical development, which could be selected for treatment of cancers in combination with compounds of the formula (I). Such agents fall into several major categories, namely, antibiotic-type agents, alkylating agents, anti-metabolite agents, hormonal agents, immunological agents, interferon-type agents and a category of miscellaneous agents. Alternatively, other chemotherapeutic agents, such as metallomatrix proteases (MMP) inhibitors may be used. Suitable agents which may be used in combination therapies include those listed, for example, in the Merck Index, An Encyclopaedia of Chemicals, Drugs and Biologicals, 12th Ed., 1996, the entire contents of which are incorporated herein by reference.
  • MMP metallomatrix proteases
  • compounds of the formula (I) When used in the treatment of solid tumours compounds of the formula (I) may be administered with one or more of the following chemotherapeutic agents: adriamycin, taxol, docetaxel, fluorouracil, melphalan, cisplatin, alpha interferon, COMP (cyclophosphamide, vincristine, methotrexate and prednisone), etoposide, mBACOD (methotrexate, bleomycin, doxorubicin, cyclophosphamide, vincristine and dexamethasone), PROMACE/MOPP (prednisone, methotrexate (w/leucovin rescue), doxorubicin, cyclophosphamide, taxol, etoposide/mechlorethamine, vincristine, prednisone and procarbazine), vincristine, vinblastine, angioinhibins, TNP 470
  • compositions comprising compounds of the formula (I) and vincristine or paclitaxel are synergistic. Accordingly, the present invention also relates to a combination comprising a compound of the general formula (I) and vincristine or paclitaxel.
  • the present invention further relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of the general formula (I) vincristine or paclitaxel, and a pharmaceutically acceptable carrier, diluent or excipient.
  • the present invention further relates to a kit of parts comprising a pharmaceutical composition of a compound of the general formula (I) and a pharmaceutical composition of vincristine or paclitaxel, wherein the compositions are intended for simultaneous, concurrent, separate or sequential use.
  • the kit may further comprise instructions for administration of the compound of the general formula (I) and vincristine or paclitaxel to a subject.
  • the present invention further relates to a method for the treatment of cancer in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) and a therapeutically effective amount of vincristine or paclitaxel.
  • the cancer may be any cancer as described herein.
  • Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential, as described above.
  • the present invention further relates to use of a compound of formula (I) in the manufacture of a medicament for treating cancer, wherein the medicament is intended for administration with vincristine or paclitaxel.
  • the cancer may be any cancer described herein.
  • Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential, as described above.
  • Tosyl hydrazide (4.35 g, 23.3 mmol) was added to a stirred solution of 2,3-dimethyl-lH-indole-5-carbaldehyde (2.7 g, 16 mmol) in dry 1,4-dioxane (50 mL) at room temperature. The temperature was increased to 80 °C and maintained for 3 hours. The crude N'-((2,3-dimethyl-lH-indol-5-yl)methylene)-4- methylbenzenesulfonohydrazide in the reaction mixture was used without isolation or further purification.
  • Tosyl hydrazide (663 mg, 3.5 mmol) was added to a stirred solution of 2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole-5-carbaldehyde (778 mg, 2.38 mmol) in dry 1,4-dioxane (30 mL) at room temperature, followed by the addition of glacial acetic acid (0.2 mL, cat.). The mixture was stirred at room temperature for 3 hours.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel using 5-10% methanol in dichloromethane as an eluent, to afford N-(3,5- dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide as a brown gummy oil (300 mg, 67%).
  • reaction mixture was quenched with saturated NaHC0 3 , then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to obtain the crude product.
  • Tosyl hydrazide (2.3 g, 12 mmol) was added to a stirred solution of 2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole-5-carbaldehyde (2.7 g, 8.2 mmol) in dry 1,4-dioxane (60 mL) at room temperature, followed by the addition of glacial acetic acid (0.7 mL, cat.). The mixture was stirred at room temperature for 3 hours. To this solution was added K 2 CO 3 (4.5 g, 33 mmol) and (4-(methoxycarbonyl)phenyl)boronic acid (2.2 g, 12 mmol).
  • the reaction temperature was raised to 110 °C and maintained for 5 hours. After complete consumption of the starting material, the reaction mass was concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using 10% methanol in dichloromethane as an eluent, to obtain methyl 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoate as a yellow solid (1.4 g, 38%).
  • LCMS m/z 448.5 [M+H] + .
  • BBr 3 (142 mL, 1.0 M in DCM) was added to a stirred solution of 5-methoxy- 2,3-dimethyl-lH-indole (10.0 g, 57.1 mmol) in dichloromethane (200 mL) at 0 °C. The temperature was maintained at 0-5 °C for 3 hours. After complete consumption of the starting material, the reaction mixture was basified with saturated NaHC0 3 then extracted with dichloromethane. The organic layer was washed with brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the organic layer was washed with water and brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using 13% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-hydroxy-2,3-dimethyl- lH-indole- 1-carboxylate as a yellow solid (11.2 g, 84%).
  • the filtrate was diluted with water and extracted with dichloromethane.
  • the organic layer was washed with brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using 3% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-(3-(methoxycarbonyl)phenoxy)-2,3-dimethyl- lH-indole-1 -carboxylate as a brown liquid (3.5 g, 46%).
  • the organic layer was washed with chilled ammonium chloride solution and brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude product was purified by flash column chromatography, using 4% ethyl acetate in petroleum ether as an eluent, to afford methyl 3-((l-(3-chloropropyl)-2,3-dimethyl- lH-indol-5-yl)oxy)benzoate as a yellow liquid (600 mg, 20%).
  • the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by flash column chromatography, using 5% methanol in dichloromethane as a eluent, to afford methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzoate (380 mg, 44%).
  • the organic layer was washed with water and brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using ethyl acetate and 5% methanol in dichloromethane as an eluent, to afford N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- 1-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide as a brown solid (50 mg, 34%).
  • the organic layer was washed with chilled ammonium chloride solution, then brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compounds was purified by column chromatography on 100-200 mesh silica gel using petroleum ether as an eluent, to afford 5-bromo-l-(3-chloro-2- methylpropyl)-2,3-dimethyl-lH-indole as a yellow liquid (6 g, crude).
  • the filtrate was diluted with water and extracted with dichloromethane.
  • the organic layer was washed with brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using 10% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-(4-(methoxycarbonyl)phenoxy)-2,3-dimethyl- lH-indole-1 -carboxylate as an off white solid (6.0 g, 66%).
  • the organic layer was washed with water and brine, then dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to obtain the crude product.
  • the crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as an eluent, to afford N-(3,5-dimethoxy-4-(pyridin- 4-yl)phenyl)-4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide as a brown solid (100 mg, 44%).
  • the organic layer was washed with brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to give the crude product.
  • the crude compound was purified by flash column chromatography using 5% ethyl acetate in petroleum ether as an eluent to afford 5-bromo- l-(3-chloropropyl)-2,3-dimethyl-lH- indole as a pink solid (2.6 g, 39%).
  • reaction mixture was heated to 100 °C for 12 hours in a sealed tube. After complete consumption of the starting material, the reaction mixture was diluted with ethyl acetate and filtered through Celite. The organic layer was washed with water and brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by prep-TLC using 5% methanol in dichloromethane as an eluent to afford N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol- 5-yl)amino)benzamide as a brown solid (80 mg, 23%).
  • reaction mixture was heated to 100 °C for 12 hours in a sealed tube. After complete consumption of the starting material, the reaction mixture was diluted with ethyl acetate and filtered through a plug of Celite. The organic layer was washed with brine solution, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to afford the crude product.
  • the crude compound was purified by prep-TLC, using 5% methanol in dichloromethane as an eluent to afford 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide as a brown gummy solid (120 mg, 34%).
  • reaction mixture was quenched with saturated NaHC0 3 , then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with brine, dried over anhydrous Na 2 S0 4 and concentrated under reduced pressure to obtain the crude product.
  • SK-N-SH neuroblastoma cells were seeded at 30 x 10 cells/well in a in a volume of 1000 ⁇ ⁇ with complete media onto a 12 well plate containing 19 mm coverslips and left to plate down for 24 hours prior to treatment. Cells were then treated with 0, 2.5 and 5 ⁇ of the test compounds.
  • each cell line was then exposed to various concentrations of each respective analogue (30, 10, 3, 1, 0.3 and 0.1 ⁇ ), cultured for a further 72 h and exposed to cell-titre luminescent reagent (100 ⁇ / ⁇ ) for a further 30 min.
  • Luminescence was captured using an En Vision multilabel reader and the data for each analogue concentration compared against no treatment control.
  • Cell viability was normalized to control (vehicle alone) and dose-response curves, and half maximal effective concentration (EC 50 ) values were determined using Graph Pad Prism 6 (nonlinear regression sigmoidal dose-response variable slope).
  • SHEP and SK-Mel-28 cell lines were maintained as monolayers in Dulbeccos Modified Eagles medium (DMEM-Invitrogen) supplemented with 10% fetal bovine serum (FBS) and grown at 37 °C in a humidified atmosphere with 5% C0 2 .
  • DMEM-Invitrogen Dulbeccos Modified Eagles medium
  • FBS fetal bovine serum
  • 2 x 10 J cells were plated into 96 well plates and treated with serial dilutions of drug (1:2 starting concentration of 200 ⁇ ) and viability measured after 48 hours using a standard MTS assay.
  • Stock solutions (10 mM) were prepared by dissolving compounds in 100% sterile DMSO. Vincristine and paclitaxel were further diluted to 0.1 mM for experiments. Small aliquots (20 ⁇ ) were prepared and stored at -30 °C under minimum light. Stock solutions were defrosted once only for each experiment.
  • the A549 and A375 cells were cultured in DMEM cell culture medium supplemented with 10% (v/v) FBS and the DU145 cells were cultured in RPMI cell culture medium supplemented with 10% (v/v) FBS. All cell lines were cultured in the absence of antibiotics for no more than 10 passages. The cells were harvested at -80% confluence by trypsinisation (passage +111 (A549), passage +20 (A375) and passage 8 (DU145)). Cells were then washed, counted and plated in 384-well plates.
  • test compounds positive control (20 ⁇ Thonzonium bromide, 100% kill) and vehicle only (0.4% DMSO) were added into wells of each assay plate using a Tecan HP D300 Digital Dispenser in technical triplicate under minimum light.
  • metabolic activity was detected by addition of 10% (v/v) alamar blue reagent to each assay well using a Multidrop Combi (Thermo Scientific) and determined by measurement of fluorescence intensity (excitation 555 nm, emission 585 nm) using an EnSpire (Perkin Elmer) plate reader. Readings were performed at 0 hour (background) and after 6 hours alamar blue incubation at 37 °C.
  • IC 50 concentrations of the compounds together with vincristine and paclitaxel as single agents against all cell lines were determined for a 72 h drug exposure using the growth inhibition assay with alamar blue readout in triplicate in two independent experiments. Each drug was tested as a 10-point concentration series with two-fold serial dilutions of each drug. The concentration series for the compounds for all cell lines tested was 20, 10, 5, 2.5, 1.25, 0.625, 0.3125, 0.156, 0.078 and 0.039 ⁇ for the two independent experiments. The concentration series for vincristine and paclitaxel was 100, 50, 25, 12.5, 6.25, 3.125, 1.5625, 0.781, 0.391 and 0.195 nM.
  • Thonzonium bromide (20 ⁇ ) and 0.4% DMSO were used as positive and vehicle controls, respectively.
  • the IC50 value was calculated by derivation of the best-fit line using a sigmoidal dose response model (Activity Base Software Suite, IDBS). Drug combination screen
  • Each drug combination was tested against A375, A549 and DU145 cells using a 6x6 dose matrix with 2-fold dilution steps based on the calculated IC50 values using the growth inhibition assay with 72h drug exposure in a 384-well format.
  • the two drugs in each combination matrix were dispensed using a Tecan HP D300 Digital Dispenser.
  • the layout of each screening plate is shown in Figure 3.
  • Assays were performed in triplicate for each drug combination (ATM/vincristine and ATM/paclitaxel combinations). Synergistic drug interactions were calculated from the averaged cell viability data using the methodology described below.
  • IDBS Activity Base software suite
  • IC 50 concentrations were calculated by derivation of the best-fit line using a sigmoidal dose response model with technical triplicate data points.
  • the Bliss additivism model was used to calculate a predicted combined response C to two single agents with responses A and B (using GI values), as follows:
  • the IC 50 concentrations of the compounds, vincristine and paclitaxel as single agents were determined against A375, A549 and DU145 cells using the growth inhibition assay with alamar blue cell viability readout.
  • the data is listed as IC 50 mean from technical triplicates for two independent experiments in Table 3.
  • the averaged IC 50 concentrations for the single agents from both runs were then used to generate the 6x6 dose matrices in the synergy experiment.
  • the compounds were tested in combination with vincristine and paclitaxel as a 6x6 dose matrix in triplicate for each cell line.
  • Synergy scores for each drug combination were calculated by the Bliss-independence model.
  • the max synergy and total synergy scores for each combination in all cell lines tested are listed in Tables 4- 6 below.
  • Max synergy is the highest individual synergy score for each combination matrix
  • Total synergy is the summation of each individual synergy score for the matrix.

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Abstract

The present invention relates broadly to functionalised and substituted indoles, the preparation thereof and their use in the treatment of proliferative diseases such as cancer.

Description

Functionalised and substituted indoles as anti-cancer agents
Field of the invention
The present invention relates broadly to functionalised and substituted indoles, the preparation thereof and their use in the treatment of proliferative diseases such as cancer.
Background of the invention
Cancer kills many thousands of people and is the second largest cause of death in the USA. There have been significant breakthroughs made in treating or preventing a wide variety of cancers. For example patients with breast cancer have benefited from early screening programs as well as a variety of surgical techniques. However, these often prove physically and emotionally debilitating. Moreover, patients who have undergone surgery and subsequent chemotherapy often experience a recurrence in their disease.
A potential new method of specifically attacking cancer cells is through disruption of cancer cells' cellular skeletal system which is comprised predominantly of actin. The actin cytoskeleton is intimately involved in cell division and cell migration. However, actin plays a ubiquitous role as the cytoskeleton of tumor cells and the actin filaments of the muscle sarcomere. The differing roles but similarity in structure make actin a difficult target for drug development due to unwanted off- target side effects.
The invention seeks to address one or more of the above mentioned problems, and/or to provide improvements in cancer therapy.
Summary of the invention
In a first aspect the present invention provides a compound of the formula (I),
Figure imgf000003_0001
(I) or a pharmaceutically acceptable salt, hydrate, derivative, solvate or prodrug thereof, wherein:
R 1 and R 2" are independently selected from the group consisting of hydrogen and Ci- C6 alkyl;
R 3J is NR 8°R 110U or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6 and wherein the ring is optionally substituted with R ;
R4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6;
Figure imgf000003_0002
each Y is independently OH or Ci-C6 alkoxy and X is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with between 1 and 3 substituents selected from the group consisting of: halo and Ci-C6 alkyl, or
R5 is
Figure imgf000003_0003
wherein Z is OH or Ci-C6 alkoxy and Q is selected from the group consisting of: halo, -S02Ci-C6 alkyl, -(CH2)o-5COOH, -(CH^o-sCOOQ-Ce alkyl or a 5- or 6-membered carbocyclic ring wherein between 1 and 4 carbons may optionally be replaced by N or NR6;
R6 is selected from the group consisting of: H and Ci-C6 alkyl; R is selected from the group consisting of: H, halo, Ci-C6 alkyl, Ci-C6alkoxy, CN, CF3 and OCF3;
R 8° and R 110U are independently selected from the group consisting of: H and Ci-C6 alkyl;
X1 is an alkanediyl group having between 1 and 20 carbon atoms; and
X 2" and 3
X3 are independently selected from the group consisting of: -0-, -NH-, -NCi- C6alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- or an alkanediyl group having between 1 and 20 carbon atoms wherein between 1 and 3 hydrogen atoms may optionally be replaced with R .
In a second aspect the present invention provides a pharmaceutical composition comprising a compound of formula (I) according to the first aspect together with a pharmaceutically acceptable carrier, diluent or excipient.
In a third aspect the present invention provides a method for the treatment of a proliferative disease in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) according to the first aspect, or a pharmaceutical composition according to the second aspect.
In a fourth aspect the present invention provides use of a compound of formula (I) according to the first aspect in the manufacture of a medicament for the treatment of a proliferative disease.
In a fifth aspect the present invention provides use of a compound of formula (I) according to the first aspect or a pharmaceutical composition according to the second aspect for the treatment of a proliferative disease in a subject.
In a sixth aspect the present invention provides a compound of formula (I) according to the first aspect or a pharmaceutical composition according to the second aspect for use in the treatment of a proliferative disease in a subject. The proliferative disease may be cancer, such as for example a solid tumour. In some embodiments the cancer is selected from the group consisting of: breast cancer, lung cancer, prostate cancer, ovarian cancer, uterine cancer, brain cancer, skin cancer, colon cancer, bladder cancer, melanoma and neuroblastoma.
The cancer may be a cancer that has recurred.
In a seventh aspect the present invention provides a method for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence, the method comprising administration to the subject of an effective amount of a compound of formula (I) according to the first aspect, or a pharmaceutical composition according to the second aspect.
In an eighth aspect the present invention provides use of a compound of formula (I) according to the first aspect in the manufacture of a medicament for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
In a ninth aspect the present invention provides a compound of formula (I) according to the first aspect for use in reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
In a tenth aspect the present invention provides use of a compound of formula (I) according to the first aspect for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
The subject may be a subject who is in cancer remission.
In an eleventh aspect the present invention provides a combination comprising:
(i) a compound of the general formula (I) according to the first aspect, and,
(ii) vincristine or paclitaxel.
The combination may be a synergistic combination. In a twelfth aspect the present invention provides a pharmaceutical composition comprising:
(i) a compound of the general formula (I) according to the first aspect,
(ii) vincristine or paclitaxel, and
(iii) a pharmaceutically acceptable carrier, diluent or excipient. The composition may be a synergistic composition.
In a thirteenth aspect the present invention provides a kit comprising:
(i) a pharmaceutical composition of a compound of the general formula (I) according to the first aspect,
and
(ii) a pharmaceutical composition of vincristine or paclitaxel, wherein the compositions are intended for simultaneous, concurrent, separate or sequential use.
In a fourteenth aspect the present invention provides a method for the treatment of cancer in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) according to the first aspect, and a therapeutically effective amount of vincristine or paclitaxel.
The cancer may be melanoma, lung cancer or prostate cancer.
Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential.
In the eleventh to fourteenth aspects the compound of formula (I) may be selected from one or more of compounds 6, 8, 14, 16, 23, 24, 27, 31, 32, 38, 39, 47, 48, 51, 55, 58, 60, 63, 67 and 68.
Further aspects of the present invention and further embodiments of the aspects described in the preceding paragraphs will become apparent from the following description, given by way of example and with reference to the accompanying drawings. Definitions
The following are some definitions that may be helpful in understanding the description of the present invention. These are intended as general definitions and should in no way limit the scope of the present invention to those terms alone, but are put forth for a better understanding of the following description.
The terms "a" and "an" are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.
Throughout this specification, unless the context requires otherwise, the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps. Thus, in the context of this specification, the term "comprising" means "including principally, but not necessarily solely".
The term "alkyl" is taken to mean straight chain or branched chain monovalent saturated hydrocarbon groups having the recited number of carbon atoms. Examples of alkyl groups include, but are not limited to, methyl, ethyl, 1 -propyl, isopropyl, 1- butyl, 2-butyl, isobutyl, tert-butyl, amyl, 1,2-dimethylpropyl, 1,1-dimethylpropyl, pentyl, isopentyl, hexyl, 4-methylpentyl, 1-methylpentyl, 2-methylpentyl, 3- methylpentyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl, 1,2-dimethylbutyl, 1,3- dimethylbutyl, 1,2,2-trimethylpropyl, 1,1,2-trimethylpropyl, 2-ethylpentyl, 3- ethylpentyl, heptyl, 1-methylhexyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 4,4- dimethylpentyl, 1,2-dimethylpentyl, 1,3-dimethylpentyl, 1,4-dimethylpentyl, 1,2,3- trimethylbutyl, 1,1,2-trimethylbutyl, 1,1,3-trimethylbutyl, 5-methylheptyl, 1- methylheptyl, octyl, nonyl and decyl.
The term "alkoxy" is taken to mean O-alkyl groups in which alkyl is as defined herein. Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, sec-butoxy and tert-butoxy. The terms "halogen" and "halo" are synonymous and refer to fluorine, chlorine, bromine or iodine.
The term "alkanediyl" is understood to refer to a bivalent saturated branched or straight chain hydrocarbon group conforming to the formula CnH2n.
The term "carbocyclic ring" refers to a ring structure in which all of the ring members are carbon atoms.
The term "pharmaceutically acceptable salt" refers to those salts which, within the scope of sound medical judgement, are suitable for use in contact with the tissues of humans and animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
The term "prodrug" means a compound which is able to be converted in vivo by metabolic means (e.g. by hydrolysis, reduction or oxidation) to a compound of the formula (I). For example, an ester prodrug of a compound of the present invention containing a hydroxy group may be hydrolysed in vivo to the parent molecule. Suitable esters are for example, acetates, citrates, lactates, tartrates, malonates, oxalates, salicylates, propionates, succinates, fumarates, maleates, methylene-bis-β- hydroxynaphthoates, gestisates, isethionates, di-/?-toluoyltartrates, methanesulfonates, ethanesulfonates, benzenesulfonates, /?-toluenesulfonates, cyclohexylsulfamates and quinates.
The terms "treating", "treatment", "preventing" and "prevention" refer to any and all uses which remedy a proliferative disease or symptoms thereof, prevent the establishment of a proliferative disease, or otherwise prevent, hinder, retard or reverse the progression of a proliferative disease or other undesirable symptoms in any way whatsoever. Thus, the terms "treating", "treatment", "preventing" and "prevention" and the like are to be considered in their broadest context. For example, treatment does not necessarily imply that a subject is treated until total recovery.
The term "therapeutically effective amount" includes a non-toxic but sufficient amount of an active compound to provide the desired therapeutic effect. Those skilled in the art will appreciate that the exact amount of a compound required will vary based on a number of factors and thus it is not possible to specify an exact "therapeutically effective amount". However, for any given case an appropriate "therapeutically effective amount" may be determined by one of ordinary skill in the art.
The term "effective amount" includes a non-toxic but sufficient amount of an active compound to provide the stated effect. When used in reference to cancer recurrence "effective amount" means an amount of a compound of formula (I) that is required to reduce the incidence of, or risk of an individual experiencing cancer recurrence. Those skilled in the art will appreciate that the exact amount of a compound required will vary based on a number of factors and thus it is not possible to specify an exact "effective amount". However, for any given case an appropriate "effective amount" may be determined by one of ordinary skill in the art.
A "pharmaceutical carrier, diluent or excipient" includes, but is not limited to, any physiological buffered (i.e., about pH 7.0 to 7.4) medium comprising a suitable water-soluble organic carrier, conventional solvents, dispersion media, fillers, solid carriers, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents. Suitable water-soluble organic carriers include, but are not limited to saline, dextrose, corn oil, dimethylsulfoxide, and gelatin capsules. Other conventional additives include lactose, mannitol, corn starch, potato starch, binders such as crystalline cellulose, cellulose derivatives, acacia, gelatins, disintegrators, such as sodium carboxymethyl-cellulose, and lubricants such as talc or magnesium stearate.
The term "subject" includes any human or non-human animal. Thus, in addition to being useful for human treatment, the compounds of the present invention may also be useful for veterinary treatment of mammals, including companion animals and farm animals, such as, but not limited to dogs, cats, horses, cows, sheep, and pigs.
The term "recurrence" as it relates to cancer is understood to mean the return of cancerous cells and/or a cancerous tumour after cancerous cells and/or a cancerous tumour have been successfully treated previously. The term "administering" and variations of that term including "administer" and "administration", include contacting, applying, delivering or providing a compound or composition of the invention to an organism, or a surface by any appropriate means.
Brief description of the drawings
Figure 1: Imaging and quantitation of actin filaments in SK-N-SH neuroblastoma cells treated with compound (A) 50, (B) 51. Cells were stained with 488-Atto-Phallodin and DAPI to visualize the actin filament bundles and the nucleus, respectively. Shown in the top panel (enlarged inset bottom panel) is a representative gray scale immunofluorescent image from control (vehicle alone), 2.5 μΜ and 5 μΜ treated cells, overlayed with the linear feature quantitation. The coloured lines indicate the detected actin filaments. Also shown is the quantitation of cell number and filament number/cell. Statistical analysis was performed using a one way ANNOVA-multiple comparison where each drug-treated group was compared to the control. **** p<0.0001, **** p<0.001, *** p<0.01, ** p<0.1.
Figure 2: Imaging and quantitation of actin filaments in SK-N-SH neuroblastoma cells treated with compound (A) 50 and (B) 51. Cells were stained with j9d primary antibody (MAb culture s/n clone 2G10.2, 1 :50) followed by 488- conjugated secondary antibody (Goat anti mouse 488, 1: 1000) and DAPI to visualize the Tpm3.1 containing filament bundles and the nucleus, respectively. Shown in the top panel (enlarged inset bottom panel) is a representative gray scale immunofluorescent image from control (vehicle alone), 2.5 μΜ and 5 μΜ treated cells, overlayed with the linear feature quantitation. The coloured lines indicate the detected actin filaments. Also shown is the quantitation of cell number and filament number/cell. Statistical analysis was performed using a one way ANNOVA-multiple comparison where each drug treated group was compared to the control. **** p<0.0001, **** p<0.001, *** p<0.01, ** p<0.1.
Figure 3: Plate layout for drug combination assays. Each screening plate contained up to six 6x6 dose matrix along with assay controls (thonzonium bromide and DMSO) as indicated. The rest of the wells contained cells only. Individual drug doses within the matrix were based on the established IC50 concentrations for the two drugs using two-fold dilution steps. Each drug combination was tested in triplicate.
Detailed description
The present invention is based on the surprising finding by the inventors that compounds of the general formula (I) effectively inhibit tropomyosin which results in an unexpected improvement in the treatment of proliferative diseases, particularly cancer.
The development of the actin cytoskeleton involves a number of ancillary control and regulatory proteins. Identification and specific targeting of actin regulatory proteins associated with the cytoskeleton of cancer cells provides the opportunity to develop cancer- specific drugs lacking unwanted side effects.
Actin filaments are constructed through the polymerisation of globular actin protein monomers. The actin monomer is polar, with one end bearing a positive charge and the other end a negative charge. The actin filaments thus have all the actin proteins aligned in one direction. These filaments have secondary coiled proteins, tropomyosins, associated with them. The tropomyosins play an integral role in regulating the function of actin filaments. Structurally, the actin filaments are made up of polymeric actin monomers with tropomyosin dimers sitting in the alpha helical groove of the actin filament to form a homopolymer. There are more than 40 mammalian tropomyosin isoforms, each of which regulates specific actin filaments. There are specific isoforms of tropomyosins that regulate the cytoskeleton of cancer cells; disruption of this interaction offers a basis to treat cancer cells with a high degree of specificity.
In one aspect the present invention provides a compound of the formula (I),
Figure imgf000012_0001
(I) or a pharmaceutically acceptable salt, hydrate, derivative, solvate or prodrug thereof, wherein:
R 1 and R 2" are independently selected from the group consisting of hydrogen and Ci- C6 alkyl;
R 3J is NR 8°R 110U or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6 and wherein the ring is optionally substituted with R ;
R4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6;
Figure imgf000012_0002
each Y is independently OH or Ci-C6 alkoxy and X is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with between 1 and 3 substituents selected from the group consisting of: halo and Ci-C6 alkyl, or
R is Q wherein Z is OH or Ci-C6 alkoxy and Q is selected from the group consisting of: halo, -S02Ci-C6 alkyl, -(CH2)o-5COOH, -(CH^o-sCOOQ-Ce alkyl or a 5- or 6-membered carbocyclic ring wherein between 1 and 4 carbons may optionally be replaced by N or NR6;
R6 is selected from the group consisting of: H and Ci-C6 alkyl; R is selected from the group consisting of: H, halo, Ci-C6 alkyl, Ci-C6alkoxy, CN, CF3 and OCF3;
R 8° and R 110U are independently selected from the group consisting of: H and Ci-C6 alkyl;
X1 is an alkanediyl group having between 1 and 20 carbon atoms; and
X 2" and X 33 are independently selected from the group consisting of: -0-, -NH-, -NCi- C6alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- or an alkanediyl group having between 1 and 20 carbon atoms wherein between 1 and 3 hydrogen atoms may optionally be replaced with R .
In one embodiment R 1 and R 2 are independently selected from the group consisting of: H and Ci-C3 alkyl.
In another embodiment R 1 and R 2 are independently selected from the group consisting of: H and methyl.
In a further embodiment R 1 and R 2 are both methyl.
In a further embodiment R 3 is NR 8 R 10 or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with R .
In yet another embodiment R 3 is NR 8 R 10 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with Ci-C6 alkyl.
In still a further embodiment R 3 is NR 8 R 10 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6.
In still a further embodiment R is N(Me)2, NH2 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6. In yet another embodiment R is N(Me)2, NH2 or a 5- or 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N or NR6.
In still another embodiment R is N(Me)2, NH2 or a 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
In yet another embodiment R is N(Me)2 or a 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
In another embodiment R is N(Me)2, NH2 or a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, S, O or NR6.
In the above embodiments for R , the 4- to 7-membered carbocyclic ring and the 5- or 6- membered carbocyclic ring may be saturated or partially saturated.
In another embodiment R is selected from the group consisting of: N(Me)2,
Figure imgf000014_0001
In another embodiment R is selected from the group consisting of: N(Me)2,
Figure imgf000014_0002
odiment R is selected from the group consisting of:
Figure imgf000014_0003
N(Me)2 and
In another embodiment R4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, NR6, O or S. In a further embodiment R4 is phenylene or a 5- or 6-membered heteroarylene ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S.
In a further embodiment R4 is phenylene or a 5- or 6-membered heteroarylene ring having between 1 and 3 N heteroatoms.
In yet another embodiment R4 is phenylene or a 6-membered heteroarylene ring having between 1 and 3 N heteroatoms.
In yet another embodiment R4 is phenylene or pyridylene.
In yet another embodiment R4 is phenylene.
In still a further embodiment R4 is para-phenylene or meta-phenylene.
In one embodiment, R 5 is
Figure imgf000015_0001
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S,and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 substituents selected from the group consisting of: halo, methyl, ethyl, propyl and isopropyl, or
R is Q wherein Z is OH or OMe and Q is selected from the group consisting of: halo, -S02Me, -(CH2)0-5COOH, -(CH2)0-5COOCi-C6alkyl, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
In another embodiment, R5 is
Figure imgf000015_0002
Y is OH or OMe and X is or a 5- or 6- membered heteroaryl ring having between 1 and 3 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 halo substituents, or
R5 is
Figure imgf000016_0001
wherein Z is OH or OMe and Q is selected from the group consisting of: halo, -S02Me, -(CH2)i-5COOH, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
In a further embodiment, R5 is
Figure imgf000016_0002
Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a halo substituent, or
R5 is
Figure imgf000016_0003
wherein Z is OH or OMe and Q is selected from the group consisting of: I, -S02Me, -(CH2)i_3COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms.
In yet another embodiment R 5 is
Figure imgf000016_0004
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or
R is Q wherein Z is OH and Q is selected from the group consisting of: I, -S02Me, -(CH2)2COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms. In still a further embodiment R is
Figure imgf000017_0001
Y is OH or OMe and
X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or
R is Q wherein Z is OH and Q is selected from the group consisting of: I, -S02Me, -(CH2)2COOH and tetrazolyl.
In still a further embodiment X is selected from the group consisting of:
Figure imgf000017_0002
In yet another embodiment X is selected from the group consisting of:
In one embodime
In some embodime
Figure imgf000017_0003
nts R5 is In other embodiments R5 is
In further embodiments R5 i
In further embodiments R5 is
Figure imgf000018_0001
In some embodiments R6 is selected from the group consisting of H, methyl and ethyl.
In other embodiments R6 is selected from the group consisting of H and methyl.
In one embodiment X1 is an alkanediyl group having between 1 and 15 carbon atoms.
In another embodiment X1 is an alkanediyl group having between 1 and 10 carbon atoms.
In a further embodiment X1 is an alkanediyl group having between 1 and 6 carbon atoms.
In yet another embodiment X1 is - (CH2)1-6- or -CH2-CH(Me)-(CH2)-. In yet another embodiment X1 is - (CH2)2_5- or -CH2-CH(Me)-(CH2)-.
In one embodiment X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NCi-C6alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 15 carbon atoms. In another embodiment X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 10 carbon atoms.
In another embodiment X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 6 carbon atoms.
In yet another embodiment X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -C(0)NH-, -NHC(O)-, -CH2CH2CH2-, CH2CH2- and - CH2-.
In yet another embodiment X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -C(0)NH-, -NHC(O)- and CH2.
In a further embodiment X is selected from the group consisting of: -0-, -NH- and -CH2-.
In still a further embodiment X3 is -C(0)NH- or -NHC(O)-. Exemplary compounds of the formula (I) include the following:
Figure imgf000020_0001
Figure imgf000021_0001
15 16
Figure imgf000022_0001
Figure imgf000023_0001
31 32
Figure imgf000024_0001
Figure imgf000025_0001
Figure imgf000025_0002
Figure imgf000025_0003
Figure imgf000025_0004
47
48
Figure imgf000026_0001
55 56
Figure imgf000027_0001
Figure imgf000028_0001
Figure imgf000028_0002
Figure imgf000028_0003
Figure imgf000028_0004
In one embodiment, the compounds are:
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-3 -((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)methyl)benzamide 3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy-[l,r-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
3- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5- yl)methyl)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)methyl)benzamide
4- ((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5- yl)methyl)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)oxy)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-3 -((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)oxy)benzamide
3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)- N-(4'-fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)oxy)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)oxy)benzamide 4-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
N-(2,6-dihydroxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)- N-(4'-fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)amino)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-3 -((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)amino)benzamide
3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide 3- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)amino)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)amino)benzamide
4- ((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide
4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)benzamide
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(3-(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide
N-(2,6-dimethoxy- [1, 1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)methyl)benzamide 4-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dimethoxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5- yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(2-(4- methylpiperazin- l-yl)ethyl)- lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1 -(2-(4-methylpiperazin- l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(2-(4-methylpiperazin- 1 - yl)ethyl)- lH-indol-5-yl)methyl)benzamide
4-((2,3-dimethyl-l-(2-(4-methylpiperazin- l-yl)ethyl)- lH-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(4-(4- methylpiperazin- l-yl)butyl)-lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1 -(4-(4-methylpiperazin- l-yl)butyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(4-(4-methylpiperazin- 1 - yl)butyl)-lH-indol-5-yl)methyl)benzamide 4-((2,3-dimethyl -(4-(4-methylpiperazin- l-yl)butyl) H-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3-dimethyl- l-(5-(4- methylpiperazin- l-yl)pentyl)- lH-indol-5-yl)methyl)benzamide
N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1 -(5-(4-methylpiperazin-
1- yl)pentyl)-lH-indol-5-yl)methyl)benzamide
N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(5-(4-methylpiperazin- 1 - yl)pentyl)- lH-indol-5-yl)methyl)benzamide
4-((2,3-dimethyl-l-(5-(4-methylpiperazin- l-yl)pentyl)- lH-indol-5-yl)methyl)-N-(4'- fluoro-2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)benzamide
4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3-hydroxy-4- iodophenyl)benzamide
4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3-hydroxy-4- (methylsulfonyl)phenyl)benzamide
3- (4-(4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl- lH-indol-5-yl)oxy)benzamido)-
2- hydroxyphenyl)propanoic acid
4- ((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3-hydroxy-4- (2H-tetrazol-5-yl)phenyl)benzamide
In one embodiment the compound of formula (I) is selected from the group consisting of: compounds 9-12, 17-20, 22-24, 30-32, 44, 46-52 and 69-72.
In another embodiment the compound of formula (I) is selected from the group consisting of: compounds 1-12, 14-20, 26-28, 33-36, 38-40, 42-44 and 46-72.
In yet another embodiment the compound of formula (I) is selected from the group consisting of: compounds 22-24 and 30-32. In yet another embodiment the compound of formula (I) is selected from the group consisting of: compounds 6, 8, 16, 23, 24, 47, 48, 51, 60 and 68.
The compounds of formula (I) may include one or more chiral centres. The present invention includes all enantiomers and diastereoisomers as well as mixtures thereof in any proportions. The invention also extends to isolated enantiomers or pairs of enantiomers. Methods of separating enantiomers and diastereoisomers are well known to persons skilled in the art. In some embodiments compounds of the formula (I) are racemic mixtures. In other embodiments compounds of the formula (I) are present in optically pure form.
Compounds of the formula (I) are also taken to include hydrates and solvates. Solvates are complexes formed by association of molecules of a solvent with a compound of the formula (I). In the case of compounds of the formula (I) that are solids, it will be understood by those skilled in the art that such compounds may exist in different crystalline or polymorphic forms, all of which are intended to be within the scope of the present invention.
The compounds of formula (I) may be in the form of pharmaceutically acceptable salts. Such salts are well known to those skilled in the art. S. M. Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66: 1-19. Pharmaceutically acceptable salts can be prepared in situ during the final isolation and purification of compounds of the formula (I), or separately by reacting the free base compound with a suitable organic acid. Suitable pharmaceutically acceptable acid addition salts of the compounds of the present invention may be prepared from an inorganic acid or from an organic acid. Examples of such inorganic acids are hydrochloric, hydrobromic, hydroiodic, nitric, carbonic, sulfuric, and phosphoric acid. Appropriate organic acids may be selected from aliphatic, cycloaliphatic, aromatic, heterocyclic carboxylic and sulfonic classes of organic acids, examples of which are formic, acetic, propionic, succinic, glycolic, gluconic, lactic, malic, tartaric, citric, ascorbic, glucoronic, fumaric, maleic, pyruvic, alkyl sulfonic, arylsulfonic, aspartic, glutamic, benzoic, anthranilic, mesylic, salicylic, p-hydroxybenzoic, phenylacetic, mandelic, ambonic, pamoic, pantothenic, sulfanilic, cyclohexylaminosulfonic, stearic, algenic, β-hydroxybutyric, galactaric, and galacturonic acids. Suitable pharmaceutically acceptable base addition salts of the compounds of the present invention include metallic salts made from lithium, sodium, potassium, magnesium, calcium, aluminium, and zinc, and organic salts made from organic bases such as choline, diethanolamine, morpholine. Alternatively, organic salts made from Ν,Ν'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N-methylglucamine), procaine, ammonium salts, quaternary salts such as tetramethylammonium salt, amino acid addition salts such as salts with glycine and arginine.
The compounds of formula (I) also extend to include all derivatives with physiologically cleavable leaving groups that can be cleaved in vivo to provide the compounds of the formula (I). Suitable leaving groups include acyl, phosphate, sulfate, sulfonate, and preferably are mono-, di- and per-acyl oxy-substituted compounds, where one or more of the pendant hydroxy groups are protected by an acyl group, preferably an acetyl group. Typically, acyloxy substituted compounds are readily cleavable to the corresponding hydroxy-substituted compounds.
Compounds of the formula (I), or salts, hydrates or solvates thereof may be prepared utilising synthetic methods known to those skilled in the art. Generally, compounds of the formula (I) may be synthesised from the following 5-substituted indoles:
R R R
Figure imgf000036_0001
(II) (VIII) (XIV)
Compounds of the formula (II) provide access to compounds of the formula (I) where X 2 is C¾ and X 3 is -C(0)NH- (referred to as compounds of formula (la)) as shown below in Scheme 1.
Scheme 1 Synthesis of compounds of the formula (la)
Figure imgf000037_0001
Figure imgf000037_0002
(la)
As an alternative, compound (III) could be hydrolysed to give the free acid and subsequently reacted with R5NH2 prior to functionalisation of the 1 -position of the indole to introduce the X 1 R 3 substituent. This alternative synthetic route is shown in Scheme 2 below.
Scheme 2 Alternative synthesis of compounds of the formula (la)
Figure imgf000037_0003
R3
(la) Compounds of the formula (VIII) provide access to compounds of the formula (I) where X 2 is O and X 3 is -C(0)NH- (referred to as compounds of the formula (lb)) as shown below in Scheme 3.
Scheme 3 Synthesis of compounds of the formula (lb)
Figure imgf000038_0001
As an alternative, compound (IX) could be hydrolysed to give the free acid and subsequently reacted with R5NH2 prior to functionalisation of the 1 -position of the indole to introduce the X 1 R 3 substituent. This alternative synthetic route is shown in Scheme 4 below.
Scheme 4 Alternative synthesis of compounds of the formula (lb)
Figure imgf000039_0001
(VI II) (IX)
Figure imgf000039_0002
(xi i) ( ii i)
Figure imgf000039_0003
Compounds of the formula (XIV) provide access to compounds of the formula (I) where X is NH (referred to as compounds of formula (Ic)) as shown below in Scheme 5.
Scheme 5 Synthesis of compounds of the formula (Ic)
Figure imgf000039_0004
(XIV) (XV) (Ic)
The methods described above in Schemes 1-5 may offer one or more advantages including high yields, control of stereochemistry, minimal synthetic steps and reaction conditions that are amenable to large scale manufacture.
The methods described above are merely representative and routine modifications and variations that would be apparent to persons skilled in the art fall within the broad scope and ambit of the invention. Alternative methods for preparing compounds of the formula (I) will become evident to those skilled in the art of synthetic organic chemistry.
Compounds of the formula (I) may be used in the treatment or prevention of proliferative diseases such as cancer. The compounds and pharmaceutical compositions of the invention may be useful for the treatment of a wide variety of cancers, including but not limited to, solid tumours such as breast cancer, lung cancer (NSCLC and SCLC), prostate cancer, ovarian cancer, uterine cancer, peritoneal cancer, brain cancer (including, for example, gliomas such as glioblastoma, Diffuse Intrinsic Pontine Glioma (DIPG) and medulloblastoma), skin cancer, colon cancer, bladder cancer, colorectal cancer, gastric cancer, liver cancer, pancreatic cancer, head and neck cancer, melanoma, malignant ascites, mesothelioma and neuroblastoma. In one embodiment the cancer is selected from the group consisting of: prostate cancer, lung cancer, breast cancer, colorectal cancer, melanoma and neuroblastoma. In alternative embodiments the cancer may be a leukaemia.
Compounds of the formula (I) may also find use in treating cancer that has recurred and in reducing the incidence of, or the risk of, recurrence of cancer in a subject deemed to be at risk of cancer recurrence, for example a subject who is in cancer remission. The subject may be in remission from a solid tumour as defined herein.
Advantageously, compounds of the formula (I) may possess superior pharmaceutical properties, such as improved resistance to conjugation via glucuronyl transferases and other water-solubilising transferases such as sulfases, which may be over-expressed on proliferative cells, such as cancer cells. This may advantageously confer superior pharmaceutical properties, such as an enhanced pharmacokinetic profile through reduced conjugation and elimination.
Those skilled in the art will recognise that compounds and pharmaceutical compositions of the present invention may be administered via any route which delivers an effective amount of the compounds to the tissue or site to be treated. In general, the compounds and compositions may be administered by the parenteral (for example intravenous, intraspinal, subcutaneous or intramuscular), oral or topical route. Administration may be systemic, regional or local. In one embodiment administration may be rectal.
The particular route of administration to be used in any given circumstance will depend on a number of factors, including the nature of the cancer to be treated, the severity and extent of the cancer, the required dosage of the particular compound to be delivered and the potential side-effects of the compound.
In general, suitable compositions may be prepared according to methods that are known to those of ordinary skill in the art and may include pharmaceutically acceptable carriers, diluents and/or excipients. The carriers, diluents and excipients must be "acceptable" in terms of being compatible with the other ingredients of the composition, and not deleterious to the recipient thereof.
Examples of pharmaceutically acceptable carriers or diluents are demineralised or distilled water; saline solution; vegetable based oils such as peanut oil, safflower oil, olive oil, cottonseed oil, maize oil or coconut oil; silicone oils, including polysiloxanes, such as methyl polysiloxane, phenyl polysiloxane and methylphenyl polysiloxane; volatile silicones; mineral oils such as liquid paraffin, soft paraffin or squalane; cellulose derivatives such as methyl cellulose, ethyl cellulose, carboxymethylcellulose, sodium carboxymethylcellulose or hydroxypropylmethylcellulose; Cremaphor; cyclodextrins; lower alkanols, for example ethanol or i-propanol; lower aralkanols; lower polyalkylene glycols or lower alkylene glycols, for example polyethylene glycol, polypropylene glycol, ethylene glycol, propylene glycol, 1,3-butylene glycol or glycerin; fatty acid esters such as isopropyl palmitate, isopropyl myristate or ethyl oleate; polyvinylpyrridone; agar; carrageenan; gum tragacanth or gum acacia and petroleum jelly. Typically, the carrier or carriers will form from 10% to 99.9% by weight of the compositions.
Pharmaceutical compositions of the invention may be in a form suitable for administration by injection, in the form of a formulation suitable for oral ingestion (such as capsules, tablets, caplets, elixirs, for example), in the form of an ointment, cream or lotion suitable for topical administration, in a form suitable for delivery as an eye drop, in an aerosol form suitable for administration by inhalation, such as by intranasal inhalation or oral inhalation, in a form suitable for parenteral administration, that is, subcutaneous, intramuscular or intravenous injection.
For administration as an injectable solution or suspension, non-toxic parenterally acceptable diluents or carriers can include cyclodextrins (for example Captisol®) Cremaphor, Ringer's solution, isotonic saline, phosphate buffered saline, ethanol and 1,2 propylene glycol. To aid injection and delivery, the compounds may also be added to PEG and non-PEGylated liposomes or micelles with specific targeting tags attached to PEG moieties, such as the RGD peptide or glutathione, for aiding passage across the blood brain barrier.
Some examples of suitable carriers, diluents, excipients and adjuvants for oral use include cyclodextrins, Cremaphor, peanut oil, liquid paraffin, sodium carboxymethylcellulose, methylcellulose, sodium alginate, gum acacia, gum tragacanth, dextrose, sucrose, sorbitol, mannitol, gelatine and lecithin. In addition these oral formulations may contain suitable flavouring and colourings agents. When used in capsule form the capsules may be coated with compounds such as glyceryl monostearate or glyceryl distearate that delay disintegration.
Adjuvants typically include emollients, emulsifiers, thickening agents, preservatives, bactericides and buffering agents.
Solid forms for oral administration may contain binders acceptable in human and veterinary pharmaceutical practice, sweeteners, disintegrating agents, diluents, flavourings, coating agents, preservatives, lubricants and/or time delay agents. Suitable binders include gum acacia, gelatine, corn starch, gum tragacanth, sodium alginate, carboxymethylcellulose or polyethylene glycol. Suitable sweeteners include sucrose, lactose, glucose, aspartame or saccharin. Suitable disintegrating agents include corn starch, methylcellulose, polyvinylpyrrolidone, guar gum, xanthan gum, bentonite, alginic acid or agar. Suitable diluents include lactose, sorbitol, mannitol, dextrose, kaolin, cellulose, calcium carbonate, calcium silicate or dicalcium phosphate. Suitable flavouring agents include peppermint oil, oil of wintergreen, cherry, orange or raspberry flavouring. Suitable coating agents include polymers or copolymers of acrylic acid and/or methacrylic acid and/or their esters, waxes, fatty alcohols, zein, shellac or gluten. Suitable preservatives include sodium benzoate, vitamin E, alpha-tocopherol, ascorbic acid, methyl paraben, propyl paraben or sodium bisulphite. Suitable lubricants include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc. Suitable time delay agents include glyceryl monostearate or glyceryl distearate.
Liquid forms for oral administration may contain, in addition to the above agents, a liquid carrier. Suitable liquid carriers include water, oils such as olive oil, peanut oil, sesame oil, sunflower oil, safflower oil, coconut oil, liquid paraffin, ethylene glycol, propylene glycol, polyethylene glycol, ethanol, propanol, isopropanol, glycerol, fatty alcohols, triglycerides or mixtures thereof.
Suspensions for oral administration may further comprise dispersing agents and/or suspending agents. Suitable suspending agents include sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, sodium alginate or acetyl alcohol. Suitable dispersing agents include lecithin, polyoxyethylene esters of fatty acids such as stearic acid, polyoxyethylene sorbitol mono- or di-oleate, -stearate or -laurate, polyoxyethylene sorbitan mono- or di-oleate, -stearate or -laurate and the like.
Emulsions for oral administration may further comprise one or more emulsifying agents. Suitable emulsifying agents include dispersing agents as exemplified above or natural gums such as guar gum, gum acacia or gum tragacanth.
Methods for preparing parenterally administrable compositions are apparent to those skilled in the art, and are described in more detail in, for example, Remington's Pharmaceutical Science, 15th ed., Mack Publishing Company, Easton, Pa., hereby incorporated by reference herein.
Topical formulations may comprise an active ingredient together with one or more acceptable carriers, and optionally any other therapeutic ingredients. Formulations suitable for topical administration include liquid or semi-liquid preparations suitable for penetration through the skin to the site where treatment is required, such as liniments, lotions, creams, ointments or pastes, and drops suitable for administration to the eye, ear or nose.
Drops according to the present invention may comprise sterile aqueous or oily solutions or suspensions. These may be prepared by dissolving the active ingredient in an aqueous solution of a bactericidal and/or fungicidal agent and/or any other suitable preservative, and optionally including a surface active agent. The resulting solution may then be clarified by filtration, transferred to a suitable container and sterilised. Sterilisation may be achieved by autoclaving or maintaining at 90 °C to 100 °C for half an hour, or by filtration, followed by transfer to a container by an aseptic technique. Examples of bactericidal and fungicidal agents suitable for inclusion in the drops are phenylmercuric nitrate or acetate (0.002%), benzalkonium chloride (0.01%) and chlorhexidine acetate (0.01%). Suitable solvents for the preparation of an oily solution include glycerol, diluted alcohol and propylene glycol.
Lotions according to the present invention include those suitable for application to the skin or eye. An eye lotion may comprise a sterile aqueous solution optionally containing a bactericide and may be prepared by methods similar to those described above in relation to the preparation of drops. Lotions or liniments for application to the skin may also include an agent to hasten drying and to cool the skin, such as an alcohol or acetone, and/or a moisiteriser such as glycerol, or oil such as olive oil.
Creams, ointments or pastes according to the present invention are semi-solid formulations of the active ingredient for external application. They may be made by mixing the active ingredient in finely divided or powdered form, alone or in solution or suspension in an aqueous or non-aqueous fluid, with a greasy or non-greasy basis. The basis may comprise hydrocarbons such as hard, soft or liquid paraffin, glycerol, beeswax, a metallic soap; a mucilage; an oil of natural origin such as almond, corn, arachis, castor or olive oil; wool fat or its derivatives, or a fatty acid such as stearic or oleic acid together with an alcohol, such as propylene glycol or macrogols.
The composition may incorporate any suitable surfactant such as an anionic, cationic or non-ionic surfactant, such as sorbitan esters or polyoxyethylene derivatives thereof. Suspending agents such as natural gums, cellulose derivatives or inoraganic materials such as silicaceous silicas, and other ingredients such a lanolin, may also be included.
In some embodiments the compositions are administered in the form of suppositories suitable for rectal administration of the compounds of formula (I). These compositions are prepared by mixing the compound of formula (I) with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the compound of formula (I). Such materials include cocoa butter, glycerinated gelatin, hydrogenated vegetable oils, mixtures of polyethylene glycols of various molecular weights and fatty acid esters of polyethylene glycol.
The compositions may also be administered or delivered to target cells in the form of liposomes. Liposomes are generally derived from phospholipids or other lipid substances and are formed by mono- or multi-lamellar hydrated liquid crystals that are dispersed in an aqueous medium. Specific examples of liposomes used in administering or delivering a composition to target cells are synthetic cholesterol (Sigma), the phospholipid l,2-distearoyl-sn-glycero-3-phosphocholine (DSPC); Avanti Polar Lipids), the PEG lipid 3-N-[(-methoxy poly(ethylene glycol)2000)carbamoyl]-l,2-dimyrestyloxy-propylamine (PEG-cDMA), and the cationic lipid l,2-di-o-octadecenyl-3-(N,N-dimethyl)aminopropane (DODMA) or 1,2- dilinoleyloxy-3-(N,N-dimethyl)aminopropane (DLinDMA) in the molar ratios 55:20: 10: 15 or 48:20:2:30, respectively, PEG-cDMA, DODMA and DLinDMA. The liposome may be contracted from l,2-distearoyl-sn-glycero-3-phosphoethanolamine- N-[methoxy(polyethylene glycol)-2000] (DSPE PEG2000) and phosphatidylcholine derived from soy and hydrogenated between 50-100%, for example Soy PC-75 or Soy PC- 100. Differing MW PEG's may be used and covalently bound with various specific targeting agents such as glutathione, RGD peptides or other recognized liposome targeting agents. Any non-toxic, physiologically acceptable and metabolisable lipid capable of forming liposomes can be used. The compositions in liposome form may contain stablisers, preservatives, excipients and the like. The preferred lipids are the phospholipids and the phosphatidyl cholines (lecithins), both natural and synthetic. Methods to form liposomes are known in the art, and in relation to this, specific reference is made to: Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, N.Y. (1976), p. 33 et seq., the contents of which is incorporated herein by reference.
The compositions may also be administered in the form of microparticles or nanoparticles. Biodegradable microparticles formed from polyactide (PLA), polylactide-co-glycolide (PLGA), and epsilon-caprolactone (έ-caprlactone) have been extensively used as drug carriers to increase plasma half life and thereby prolong efficacy (R. Kumar, M., 2000, J. Pharm. Pharmaceut. Sci. 3(2) 234-258). Microparticles have been formulated for the delivery of a range of drug candidates including vaccines, antibiotics, and DNA. Moreover, these formulations have been developed for various delivery routes including parenteral subcutaneous injection, intravenous injection and inhalation.
The compositions may incorporate a controlled release matrix that is composed of sucrose acetate isobutyrate (SAIB) and an organic solvent or organic solvents mixture. Polymer additives may be added to the vehicle as a release modifier to further increase the viscosity and slow down the release rate. SAIB is a well known food additive. It is a very hydrophobic, fully esterified sucrose derivative, at a nominal ratio of six isobutyrate to two acetate groups. As a mixed ester, SAIB does not crystallise but exists as a clear viscous liquid. Mixing SAIB with a pharmaceutically acceptable organic solvent, such as ethanol or benzyl alcohol decreases the viscosity of the mixture sufficiently to allow for injection. An active pharmaceutical ingredient may be added to the SAIB delivery vehicle to form SAIB solution or suspension formulations. When the formulation is injected subcutaneously, the solvent differs from the matrix allowing the SAIB-drug or SAIB- drug-polymer mixtures to set up as an in situ forming depot.
For the purposes of the present invention compounds and compositions may be administered to subjects either therapeutically or preventively. In a therapeutic application compositions are administered to a patient already suffering from cancer in an amount sufficient to cure or at least partially arrest the cancer and its complications. The composition should provide a quantity of the compound or agent sufficient to effectively treat the subject.
The therapeutically effective amount for any particular subject will depend upon a variety of factors including: the cancer being treated and the severity thereof; the activity of the compound administered; the composition in which the compound is present; the age, body weight, general health, sex and diet of the subject; the time of administration; the route of administration; the rate of sequestration of the compound; the duration of the treatment; drugs used in combination or coincidental with the compound, together with other related factors well known in medicine.
One skilled in the art would be able, by routine experimentation, to determine an effective, non-toxic amount of a compound that would be required to treat or prevent a particular cancer.
Generally, an effective dosage is expected to be in the range of about 0.0001 mg to about 1000 mg per kg body weight per 24 hours; typically, about 0.001 mg to about 750 mg per kg body weight per 24 hours; about 0.01 mg to about 500 mg per kg body weight per 24 hours; about 0.1 mg to about 500 mg per kg body weight per 24 hours; about 0.1 mg to about 250 mg per kg body weight per 24 hours; about 1.0 mg to about 250 mg per kg body weight per 24 hours. More typically, an effective dose range is expected to be in the range about 1.0 mg to about 200 mg per kg body weight per 24 hours; about 1.0 mg to about 100 mg per kg body weight per 24 hours; about 1.0 mg to about 50 mg per kg body weight per 24 hours; about 1.0 mg to about 25 mg per kg body weight per 24 hours; about 5.0 mg to about 50 mg per kg body weight per 24 hours; about 5.0 mg to about 20 mg per kg body weight per 24 hours; about 5.0 mg to about 15 mg per kg body weight per 24 hours.
Alternatively, an effective dosage may be up to about 500 mg/m . Generally, an effective dosage is expected to be in the range of about 25 to about 500 mg/m , preferably about 25 to about 350 mg/m , more preferably about 25 to about 300
2 2
mg/m , still more preferably about 25 to about 250 mg/m , even more preferably about 50 to about 250 mg/m , and still even more preferably about 75 to about 150 mg/m2. Typically, in therapeutic applications, the treatment would be for the duration of the disease state.
Further, it will be apparent to one of ordinary skill in the art that the optimal quantity and spacing of individual dosages will be determined by the nature and extent of the cancer being treated, the form, route and site of administration, and the nature of the particular individual being treated. Also, such optimum conditions can be determined by conventional techniques.
The compounds of formula (I) may be used alone in the treatment of cancer, or alternatively in combination with radiotherapy and/or surgery and/or other therapeutic agents, for example chemotherapeutic agents and immunostimulatory agents, as part of a combination therapy. The compounds of formula (I) may sensitise cancer cells to other chemotherapeutic agents and/or radiotherapy.
The terms "combination therapy" and "adjunct therapy" are intended to embrace administration of multiple therapeutic agents in a sequential manner in a regimen that will provide beneficial effects and is intended to embrace administration of these agents in either a single formulation or in separate formulations.
Combination therapy may involve the active agents being administered together, sequentially, or spaced apart as appropriate in each case. Combinations of active agents including compounds of the invention may be synergistic.
The co-administration of compounds of the formula (I) with other therapeutic agent(s) may be effected by a compound of the formula (I) being in the same unit dose form as the other therapeutic agent(s), or the compound of the formula (I) and the other therapeutic agent(s) may be present in individual and discrete unit dosage forms that are administered concurrently, sequentially or separately. Separate administration refers to administration of the compound of the formula (I) independently of the other therapeutic agent(s) at different time points. It may be the case that the compound of the formula (I) and the other therapeutic agent(s) are administered such that no overlap of measurable blood levels of the compound formula (I) and the other therapeutic agent(s) are present. When administered separately, it may be preferred for the compound of the formula (I) and the the other therapeutic agent(s) to be administered by the same route of administration, although it is not necessary for this to be so. Sequential administration may be in any order as required, and may require an ongoing physiological effect of the first or initial agent to be current when the second or later agent is administered, especially where a cumulative or synergistic effect is desired.
In accordance with various embodiments of the present invention one or more compounds of formula (I) may be included in combination therapy with surgery and/or radiotherapy and/or one or more chemotherapeutic agents.
There are large numbers of chemotherapeutic agents that are currently in use, in clinical evaluation and in pre-clinical development, which could be selected for treatment of cancers in combination with compounds of the formula (I). Such agents fall into several major categories, namely, antibiotic-type agents, alkylating agents, anti-metabolite agents, hormonal agents, immunological agents, interferon-type agents and a category of miscellaneous agents. Alternatively, other chemotherapeutic agents, such as metallomatrix proteases (MMP) inhibitors may be used. Suitable agents which may be used in combination therapies include those listed, for example, in the Merck Index, An Encyclopaedia of Chemicals, Drugs and Biologicals, 12th Ed., 1996, the entire contents of which are incorporated herein by reference.
When used in the treatment of solid tumours compounds of the formula (I) may be administered with one or more of the following chemotherapeutic agents: adriamycin, taxol, docetaxel, fluorouracil, melphalan, cisplatin, alpha interferon, COMP (cyclophosphamide, vincristine, methotrexate and prednisone), etoposide, mBACOD (methotrexate, bleomycin, doxorubicin, cyclophosphamide, vincristine and dexamethasone), PROMACE/MOPP (prednisone, methotrexate (w/leucovin rescue), doxorubicin, cyclophosphamide, taxol, etoposide/mechlorethamine, vincristine, prednisone and procarbazine), vincristine, vinblastine, angioinhibins, TNP 470, pentosan polysulfate, platelet factor 4, angiostatin, LM 609, SU 101, CM 101, Techgalan, thalidomide, SP-PG and the like. The inventors have discovered that compositions comprising compounds of the formula (I) and vincristine or paclitaxel are synergistic. Accordingly, the present invention also relates to a combination comprising a compound of the general formula (I) and vincristine or paclitaxel.
The present invention further relates to a pharmaceutical composition comprising a compound of the general formula (I) vincristine or paclitaxel, and a pharmaceutically acceptable carrier, diluent or excipient.
In addition, the present invention further relates to a kit of parts comprising a pharmaceutical composition of a compound of the general formula (I) and a pharmaceutical composition of vincristine or paclitaxel, wherein the compositions are intended for simultaneous, concurrent, separate or sequential use. The kit may further comprise instructions for administration of the compound of the general formula (I) and vincristine or paclitaxel to a subject.
The present invention further relates to a method for the treatment of cancer in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) and a therapeutically effective amount of vincristine or paclitaxel. The cancer may be any cancer as described herein. Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential, as described above.
The present invention further relates to use of a compound of formula (I) in the manufacture of a medicament for treating cancer, wherein the medicament is intended for administration with vincristine or paclitaxel. The cancer may be any cancer described herein. Administration of the compound of formula (I) and vincristine or paclitaxel may be simultaneous, concurrent, separate or sequential, as described above.
Embodiments of the invention will now be described in more detail with reference to the examples which are provided for exemplification only and which should not be considered limiting on the scope of the invention in any way. Examples
Example 1 - Synthesis of compounds
Scheme 6. Preparation of Intermediate A.
Figure imgf000051_0001
Preparation of 1 -(2,6-difluoro-4-nitrophenyl)-lH-imidazole
A mixture of l,2,3-trifluoro-5-nitrobenzene (45 g, 0.25 mol), imidazole (17.2 g, 0.25 mol) and triethylamine (35.6 mL, 0.25 mol) in acetonitrile (250 mL) was heated at reflux for 20 hours. The mixture was poured into water and extracted with ethyl acetate. The organic layer was washed with water and brine, dried over Na2S04 and concentrated in vacuo to give l-(2,6-difluoro-4-nitrophenyl)-lH-imidazole (46 g, 80%).
1H NMR (400 MHz, CDC13): δ 8.05 (d, = 10.0 Hz, 1.8 Hz, 2H), 7.85 (br s, 1H), 7.30-7.26 (m, 2H). LCMS: m/z 226.1 [M+H]+.
Preparation of 3,5-difluoro-4-(lH-imidazol-l-yl)aniline
To a solution of l-(2,6-difluoro-4-nitrophenyl)-lH- imidazole (46 g, 0.20 mol) in ethanohwater (4: 1, 500 mL) at room temperature was added Fe powder (59 g, 1.1 mol) and NH4C1 (16.2 g, 0.30 mol). The mixture was heated to 80 °C for 5 hours. The reaction mixture was filtered through Celite and the ethanol was evaporated. The aqueous layer was extracted with ethyl acetate. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated to give the crude product. The crude compound was purified by flash column chromatography to give 3,5-difluoro- 4-(lH-imidazol-l-yl)aniline (38 g, 98%).
Preparation of Intermediate A, 4-( lH-imidazol-l-yl)-3,5-dimethoxyaniline A solution of 3,5-difluoro-4-(lH-imidazol-l-yl)aniline (10 g, 51 mmol) and NaOMe (55.3 g, 1.02 mol) in methanol (100 mL) was heated to 120 °C in a sealed tube for 24 hours. Once cooled, the reaction mixture was quenched with aqueous HCl and extracted with ethyl acetate. The organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated in vacuo to give the crude product. The crude compound was purified by flash column chromatography to afford 4-(lH-imidazol-l-yl)-3,5-dimethoxyaniline (Intermediate A) as a solid (5.0 g, 44%).
1H NMR (400 MHz, DMSO-d6): δ 7.39 (s, 1H), 6.93 (d, = 17.6 Hz, 2H), 5.97 (br s, 2H), 5.46 (br s, 2H), 3.61 (s, 6H).
Scheme 7. Preparation of Intermediates B, C and D.
Figure imgf000052_0001
B C D
Preparation of 4-bromo-3, 5 -dimethoxy aniline
To a solution of 3, 5 -dimethoxy aniline (10 g, 65 mmol) in dichloromethane (250 mL) was added tetrabutylammonium tribromide (31 g, 65 mmol) at -78 °C. The mixture was allowed to warm to room temperature and stirred for 5 hours. The mixture was poured into water and extracted with dichloromethane. The organic extracts were washed with water and brine, dried over anhydrous Na2S04 and concentrated to afford 4-bromo-3,5-dimethoxyaniline (3.2 g, 21%).
1H NMR (400 MHz, DMSO-d6): δ 6.97 (br s, 2H), 5.38 (br s, 2H), 3.73 (s, 6H). LCMS: m/z 232.0, 234.0 [M+H]+.
Preparation of Intermediate B, 3,5-dimethoxy-4-(pyridin-4-yl)aniline To a solution of 4-bromo-3,5-dimethoxyaniline (2.0 g, 8.6 mmol) in DMF (15 mL), Pd(PPh3)4 (5 mol%) and K3P04 (5.0 g, 24 mmol) were added at room temperature. The mixture was heated to 120 °C for 16 hours. The reaction mass was concentrated, diluted with water and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 3,5-dimethoxy-4-(pyridin-4-yl)aniline (Intermediate B) as a solid (2.0 g, quant). LCMS: m/z 231.1 [M+H]+.
Other analogues prepared by this method:
Intermediate C, 2,6-dimethoxy-[l,l'-biphenyl]-4-amine (quant.).
1H NMR (400 MHz, CDC13): δ 7.36-7.26 (m, 5H), 6.01 (s, 2H), 3.81-3.65 (m, 8H). LCMS: m/z 230.1 [M+H]+.
Intermediate D, 4'-fluoro-2,6-dimethoxy-[l,l'-biphenyl]-4-amine (quant.)
1H NMR (400 MHz, DMSO-d6): δ 7.71-7.54 (m, 2H), 7.20-7.04 (m, 4H), 5.97 (s, 2H), 5.23 (br s, 2H), 3.58 (s, 6H). LCMS: m/z 248.1 [M+H]+.
Scheme 8. Preparation of Compounds 1-4
Figure imgf000054_0001
Preparation of2,3-dimethyl-lH-indole-5-carbaldehyde
To a stirred solution of 5-bromo-2,3-dimethyl-lH- indole (5.6 g, 25 mmol) in dry THF (40 mL) was added i-BuLi (50 mL, 75 mmol, 1.5 M in THF) at -78 °C. The resulting reaction mixture was stirred at the same temperature for one hour. Then dry DMF (5.0 mL, 65 mmol) was added to the reaction mass at -78 °C. The mixture was stirred at this temperature for a further 2 hours. After complete consumption of the starting material, the reaction mass was quenched with saturated ammonium chloride solution at -40 °C and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford crude 2,3-dimethyl-lH-indole-5-carbaldehyde as a yellow solid (2.7 g, 62%), which was used in the next step without further purification. LCMS: m/z 174.1 [M+H]+. Preparation of N ( 2,3- dimethyl- IH-indol- -5 -yl)methylene )-4- methylbenzenesulfonohydrazide
Tosyl hydrazide (4.35 g, 23.3 mmol) was added to a stirred solution of 2,3-dimethyl-lH-indole-5-carbaldehyde (2.7 g, 16 mmol) in dry 1,4-dioxane (50 mL) at room temperature. The temperature was increased to 80 °C and maintained for 3 hours. The crude N'-((2,3-dimethyl-lH-indol-5-yl)methylene)-4- methylbenzenesulfonohydrazide in the reaction mixture was used without isolation or further purification.
Preparation of methyl 3-( ( 2,3-dimethyl-lH-indol-5-yl)methyl)benzoate
To the solution of crude 2 N'-((2,3-dimethyl- lH-indol-5-yl)methylene)-4- methylbenzenesulfonohydrazide in 1,4-dioxane, cooled to 0 °C, was added K2CO3 (3.2 g, 23 mmol) and (3-(methoxycarbonyl)phenyl)boronic acid (4.2 g, 23 mmol). The reaction temperature was raised to 110 °C and maintained for 5 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with water and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 5-10% ethyl acetate in petroleum ether as an eluent, to obtain methyl 3-((2,3-dimethyl- lH-indol-5- yl)methyl)benzoate as a brown oil (2.4 g, 53%).
1H NMR (400 MHz, CDC13): δ 7.94 (br s, 1H), 7.85 (d, 7 = 7.6 Hz, 1H), 7.63 (br s, 1H), 7.39 (d, 7 = 7.6 Hz, 1H), 7.31 (t, 7 = 7.6 Hz, 1H), 7.27 (br s, lH), 7.16 (d, 7 = 8.0 Hz, 1H), 6.91 (dd, 7 = 8.0 Hz, 1.2 Hz, 1H), 4.11 (s, 2H), 3.88 (s, 3H), 2.33 (s, 3H), 2.18 (s, 3H). LCMS: m/z 294.1 [M+H]+.
Preparation of 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoic acid
NaH (1.0 g, 42 mmol) was added portionwise to a stirred solution of methyl 3-((2,3-dimethyl-lH-indol-5-yl)methyl)benzoate (2.4 g, 8.2 mmol) in DMF (20 mL) at 0 °C. The mixture was stirred at this temperature for 30 minutes. To this, 1-bromo- 3-chloropropane (1.6 niL, 16 mmol) was added dropwise at 0 °C and the reaction mixture was allowed to stir at this temperature for 3 hours. After complete consumption of the starting material, the reaction mixture was quenched with saturated ammonium chloride solution, diluted with ice-cold water and then extracted with ethyl acetate. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 15% ethyl acetate in petroleum ether as an eluent, to afford 3- ((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5-yl)methyl)benzoic acid as a brown liquid (1.8 g, crude). LCMS: m/z 356.0 [M+H]+.
Preparation of methyl 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoate
To a solution of 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoic acid (1.8 g, 5.0 mmol) in methanol (20 mL) at 0 °C was added concentrated sulphuric acid (0.5 mL). The mixture was heated to 80 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was concentrated and diluted with ethyl acetate. The organic layer was washed with water and brine, dried over Na2S04 and concentrated in vacuo to give the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 5% ethyl acetate in petroleum ether as an eluent, to afford methyl 3-((l-(3- chloropropyl)-2,3-dimethyl-lH-indol-5-yl)methyl)benzoate as a yellow liquid (1.4 g, crude). LCMS: m/z 370.1 [M+H]+.
Preparation of methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)methyl)benzoate
To a stirred solution of methyl 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol- 5-yl)methyl)benzoate (1.4 g, 3.8 mmol) in acetonitrile (15 mL), sodium iodide (1.4 g, 9.3 mmol), sodium carbonate (1.2 g, 11 mmol) and then N-methylpiperazine (1.0 mL, 9.0 mmol) were added at room temperature. The reaction mixture was heated to 75 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, concentrated under reduced pressure and diluted with ethyl acetate. The resulting solution was washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 50% ethyl acetate in petroleum ether as an eluent, to afford methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoate as a brown oil (1.1 g, 67%). LCMS: m/z [M+H]+.
Preparation of 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoic acid
To a solution of methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoate (1.1 g, 2.5 mmol) in THF:H20:MeOH (4: 1: 1, 36 mL) was added LiOH.H20 (213 mg, 5.08 mmol) at 0 °C. The reaction mixture was stirred at room temperature for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, neutralised to pH~7 and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine solution, then dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoic acid (798 mg, 76%). LCMS: m/z 420.3 [M+H]+.
Preparation of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lll-indol-5-yl)methyl)benzamide
To a solution of 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoic acid (200 mg, 0.48 mmol) and Intermediate A (134 mg, 0.61 mmol) in pyridine (2 mL), was added EDC.HC1 (550 mg, 2.9 mmol). The mixture was stirred at room temperate for 10 minutes before being heated to 80 °C for 2 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with water and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, then dried and concentrated under reduced pressure to afford N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol- 5-yl)methyl)benzamide (110 mg, 37%). LCMS: m/z 621.3 [M+H]+.
Other analogues prepared by this method:
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (37%). LCMS: m/z 632.3 [M+H]+.
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (44%). LCMS: m/z 631.3 [M+H]+.
3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)- N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (38%). LCMS: m/z 649.2 [M+H]+.
Preparation of Compound 1, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)- 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide
To a stirred solution of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide (130 mg, 0.21 mmol) in dichloromethane (5 mL) at 0 °C was added boron tribromide (1.0 mL, 1 M in DCM). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform, washed with brine solution and dried over Na2S04. The organic layer was concentrated under reduced pressure and the crude compound was purified by prep-HPLC to afford N-(3,5-dihydroxy-4-(lH-imidazol-l- yl)phenyl)-3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide (Compound 1) as a brown solid (5 mg, 4%).
1H NMR (400 MHz, CD3OD): δ 7.75 (br s, 1H), 7.71-7.68 (m, 2H), 7.46-7.38 (m, 2H), 7.29-7.24 (m, 2H), 7.14 (br s, 1H), 7.07 (br s, 1H), 6.97-6.94 (m, 3H), 4.17 (t, J = 6.8 Hz, 2H), 4.13 (s, 2H), 2.71-2.40 (m, 8H), 2.39-2.32 (m, 5H), 2.19 (s, 3H), 2.15 (s, 3H), 1.95-1.79 (m, 2H). LCMS: m/z 593.3 [M+H]+.
Other analogues prepared by this method:
Compound 2, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (3 -(4-methylpiperazin- 1 -yl)propyl)- lH-indol-5 -yl)methyl)benzamide (18%).
1H NMR (400 MHz, CD3OD): δ 8.46 (d, = 4.8 Hz, 2H), 7.75 (br s, 1H), 7.70 (br d, = 7.2 Hz, 1H), 7.53 (d, = 4.8 Hz, 2H), 7.46 (br d, = 7.2 Hz, 1H), 7.40 (t, = 7.6 Hz, 1H), 7.29 (br s, 1H), 7.26 (d, = 8.4 Hz, 1H), 6.96 (br d, = 8.4 Hz, 1H), 6.89 (s, 2H), 4.18 (t, = 6.8 Hz, 2H), 4.13 (s, 2H), 3.07-2.35 (m, 16H), 2.20 (s, 3H), 1.92 (quintet, / = 6.4 Hz, 2H). LCMS: m/z 604.3 [M+H]+.
Compound 3, N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(3- (4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide (16%).
1H NMR (400 MHz, CD3OD): δ 7.74 (br s, 1H), 7.70 (br d, = 6.8 Hz, 1H), 7.46-7.39 (m, 2H), 7.37-7.32 (m, 4H), 7.28-7.23 (m, 3H), 6.96 (br d, = 8.4 Hz, 1H), 6.84 (br s, 2H), 4.18-4.13 (m, 4H), 2.80-2.32 (m, 16H), 2.19 (s, 3H), 1.91 (quintet, = 6.8 Hz, 2H). LCMS: m/z 603.3 [M+H]+.
Compound 4, 3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4-yl)benzamide (22%).
1H NMR (400 MHz, CD3OD): δ 7.74 (br s, 1H), 7.70 (br d, = 7.6 Hz, 1H), 7.64-7.30 (m, 6H), 7.29 (br s, 1H), 7.25 (d, = 8.4 Hz, 1H), 7.07 (t, = 8.8 Hz, 2H), 6.96 (br d, = 8.4 Hz, 1H), 6.84 (s, 2H), 4.17 (t, = 6.8 Hz, 2H), 4.13 (s, 2H), 2.89- 2.34 (m, 16H), 2.20 (s, 3H), 1.92 (quintet, = 7.2 Hz, 2H). LCMS: m/z 621.3 [M+H]+.
Scheme 9. Preparation of Compounds 5-8.
Figure imgf000060_0001
Preparation of5-bromo-l-(3-chloro-2-methylpropyl)-2,3-dimethyl-lH-indole
To a solution of NaH (2.8 g, 0.12 mol) in dry DMF (50 niL) was added dropwise a solution of 5-bromo-2,3-dimethyl-lH- indole (5.0 g, 22 mmol) in dry DMF (50 niL) at 0 °C. The mixture was stirred for 30 minutes. To this, l-bromo-3-chloro- 2-methylpropane (11 g, 64 mmol) in dry DMF (20 mL) was added dropwise at 0 °C. The mixture was stirred for 3 hours at room temperature. After complete consumption of the starting material, the reaction mixture was quenched with saturated ammonium chloride solution and diluted with ice-cold water. The reaction mixture was extracted with ethyl acetate. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude compound. The crude product was purified by column chromatography on 100-200 mesh silica gel, using 2% ethyl acetate in petroleum ether as an eluent, to afford 5- bromo-l-(3-chloro-2-methylpropyl)-2,3-dimethyl-lH-indole as a yellow liquid (6.8 g, crude). LCMS: m/z 314.0, 316.0 [M+H]+.
Preparation of 5-bromo-2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indole To a stirred solution of 5-bromo-l-(3-chloro-2-methylpropyl)-2,3-dimethyl- lH-indole (6.8 g, 22 mmol) in acetonitrile (50 niL), were added sodium iodide (8.1 g, 54 mmol) and sodium carbonate (6.9 g, 65 mmol) followed by N-methylpiperazine (5.4 g, 54 mmol) at room temperature. The reaction mixture was heated to 75 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, concentrated, diluted with ethyl acetate, washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 5% methanol in dichloromethane as an eluent, to afford 5-bromo-2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indole as a yellow oil (1.4 g, 17%). LCMS: m/z 379.1, 381.1 [M+H]+.
Preparation of 2, 3 -dimethyl- 1 -(2-methyl-3-(4-methylpiperazin-l -yl)propyl)- lH-indole-5-carbaldehyde
To a stirred solution of 5-bromo-2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indole (1.3 g, 3.4 mmol), in dry THF (30 mL) was added n-BuLi (4.1 mL, 6.6 mmol, 1.6 M in THF) at -78 °C. The resulting reaction mixture was stirred at this temperature for 15 minutes. Then dry DMF (1.5 mL) was added to the reaction mass at -78 °C. The mixture was stirred at this temperature for a further one hour. After complete consumption of the starting material, the reaction mass was quenched with saturated ammonium chloride solution and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 2% methanol in dichloromethane as an eluent, to afford 2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole-5-carbaldehyde as a yellow oil (890 mg, 80%).
1H NMR (400 MHz, CDC13): δ 9.87 (s, 1H), 7.88 (br s, 1H), 7.53 (dd, J = 8.4 Hz, 1.6 Hz, 1H), 7.19 (d, J = 8.4 Hz, 1H), 4.18-4.12 (m, 1H), 3.67-3.61 (m, 1H), 2.30-2.05 (m, 20H), 0.74 (d, J = 6.4 Hz, 3H). LCMS: m/z 328.2 [M+H]+. Preparation of N '-(( 2,3 -dimethyl- 1 -(2-methyl-3-(4-methylpiperazin-l - yl)propyl)-lH-indol-5-yl)methylene)-4-methylbenzenesulfonohydrazide
Tosyl hydrazide (663 mg, 3.5 mmol) was added to a stirred solution of 2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole-5-carbaldehyde (778 mg, 2.38 mmol) in dry 1,4-dioxane (30 mL) at room temperature, followed by the addition of glacial acetic acid (0.2 mL, cat.). The mixture was stirred at room temperature for 3 hours. The crude N'-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methylene)-4- methylbenzenesulfonohydrazide in the reaction mixture was used without isolation or further purification.
Preparation of methyl 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl )propyl)-lH-indol-5-yl )methyl )benzoate
To the solution of crude N'-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)methylene)-4-methylbenzenesulfonohydrazide in dry 1,4- dioxane was added K2CO3 (1.3 g, 9.4 mmol) and (3-(methoxycarbonyl)phenyl)boronic acid (642 mg, 3.6 mmol). The reaction temperature was raised to 110 °C and maintained for 5 hours. After complete consumption of the starting material, the reaction mass was concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 10% methanol in dichloromethane as an eluent, to afford methyl 3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzoate as a yellow solid (300 mg, crude). LCMS: m/z 448.5 [M+H]+.
Preparation of 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl )propyl)-lH-indol-5-yl )methyl )benzoic acid
To a solution of methyl 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoate (295 mg, 0.659 mmol) in THF:H20:MeOH (4: 1: 1, 12 mL) was added LiOH.H20 (55 mg, 1.3 mmol) at 0 °C. The reaction mixture was stirred at room temperature for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, neutralised to pH~7 and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 10% methanol in dichloromethane as an eluent, to afford 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoic acid (148 mg, 52%).
1H NMR (400 MHz, DMSO-d6): δ 7.78 (br s, 1H), 7.74 (br d, J = 7.6 Hz, 1H), 7.51 (br d, J = 7.2 Hz, 1H), 7.40 (t, = 8.0 Hz, 1H), 7.27 (br s, 1H), 7.24 (d, = 8.4 Hz, 1H), 6.90 (dd, = 8.0 Hz, 1.2 Hz, 1H), 4.17-4.11 (m, 1H), 4.06 (s, 2H), 3.84- 3.77 (m, 1H), 3.12-2.51 (m, 9H), 2.38-2.05 (m, 11H), 0.77 (d, = 6.8 Hz, 3H). LCMS: m/z 434.07 [M+H]+.
Preparation of N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide
To a solution of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoic acid (300 mg, 0.69 mmol) and Intermediate B (200 mg, 0.87 mmol) in pyridine (4 mL), was added EDC.HCl (800 mg, 4.2 mmol). The mixture was stirred at 80 °C for 3 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with water and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, then dried and concentrated under reduced pressure to give the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 5-10% methanol in dichloromethane as an eluent, to afford N-(3,5- dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide as a brown gummy oil (300 mg, 67%).
1H NMR (300 MHz, DMSO-d6): δ 10.31 (br s, 1H), 8.53 (d, = 6.0 Hz, 2H), 7.85 (br s, 1H), 7.81 (br d, = 6.9 Hz, 1H), 7.50-7.39 (m, 2H), 7.36 (s, 2H), 7.31- 7.22 (m, 4H), 6.93 (dd, = 8.1 Hz, 0.9 Hz, 1H), 4.17-4.05 (m, 7H), 3.82-3.77 (m, 1H), 3.69 (s, 6H), 2.49-2.08 (m, 16H), 0.73 (d, J = 6.3 Hz, 3H). LCMS: m/z 646.65 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3-((2,3-dimethyl-l-(2-methyl- 3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (64%). LCMS: m/z 635.56 [M+H]+.
N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (67%). LCMS: m/z 645.45 [M+H]+.
3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (69%). LCMS: m/z 663.67 [M+H]+.
Preparation of Compound 5, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)- 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide
To a stirred solution of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide (280 mg, 0.44 mmol) in dichloromethane (6 mL) at 0 °C was added boron tribromide (3.0 mL, 3.0 mmol, 1 M in DCM). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by prep-TLC using 2% methanol in dichloromethane as an eluent, to afford N-(3,5-dihydroxy-4- (lH-imidazol-l-yl)phenyl)-3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzamide (Compound 5) as an off white solid (14 mg, 5%). 1H NMR (400 MHz, DMSO-d6): δ 10.13 (br s, IH), 9.89 (br s, 2H), 7.80 (br s, IH), 7.73 (br s, 7.2 Hz, IH), 7.53 (br s, IH), 7.45-7.38 (m, 2H), 7.28 (br s, IH), 7.22 (d, = 8.0 Hz, IH), 7.09 (br s, IH), 7.02 (s, 2H), 6.95 (br s, IH), 6.92 (dd, = 8.4 Hz,
1.6 Hz, IH), 4.14 (dd, = 14.8 Hz, 5.2 Hz, IH), 4.08 (s, 2H), 3.78 (dd, = 14.8 Hz, 8.8 Hz, IH), 2.43-2.12 (m, 20H), 0.72 (d, = 6.4 Hz, 3H). LCMS: m/z 605.69 [M-H]~
Other analogues prepared by this method:
Compound 6, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (7%).
1H NMR (300 MHz, DMSO-d6): δ 10.10 (br s, IH), 9.53 (br s, 2H), 8.49 (d, = 6.3 Hz, 2H), 7.80 (br s, IH), 7.73 (br d, = 6.6 Hz, IH), 7.43-7.39 (m, 2H), 7.36 (d, = 6.0 Hz, 2H), 7.28 (br s, IH), 7.23 (d, = 8.4 Hz, IH), 6.98 (s, 2H), 6.92 (dd, =
8.7 Hz, 1.8 Hz, IH), 4.14 (dd, = 15.0 Hz, 4.8 Hz, IH), 4.08 (s, 2H), 3.78 (dd, = 15.3 Hz, 9.0 Hz, IH), 2.41-2.12 (m, 20H), 0.72 (d, = 6.3 Hz, 3H). LCMS: m/z 618.43 [M+H]+.
Compound 7, N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2- methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (5%).
1H NMR (300 MHz, DMSO-d6): δ 7.80 (br s, IH), 7.73 (br d, = 6.6 Hz, IH), 7.44-7.17 (m, 9H), 6.95-6.89 (m, 3H), 4.17-4.06 (m, 3H), 3.78 (dd, = 14.7 Hz, 8.4 Hz, IH), 2.37-2.09 (m, 20H), 0.73 (d, = 6.0 Hz, 3H). LCMS: m/z 617.67 [M+H]+.
Compound 8, 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)- lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4- yl)benzamide (22%).
1H NMR (400 MHz, CD3OD): δ 7.76 (br s, IH), 7.69 (br d, = 7.6 Hz, IH), 7.46-7.35 (m, 4H), 7.28 (br s, IH), 7.20 (d, = 8.4 Hz, IH), 7.06 (t, = 8.8 Hz, 2H), 6.95 (dd, = 8.0 Hz, 1.2 Hz, IH), 6.84 (s, 2H), 4.18-4.06 (m, 3H), 3.89-3.83 (m, IH), 2.59-2.20 (m, 20H), 0.86 (d, = 5.6 Hz, 3H). LCMS: m/z 635.56 [M+H]+. Scheme 10. Preparation of Compounds 9-12 and 49-52
Figure imgf000066_0001
49: X = 1 -imidazolyl 9: X = 1-imidazolyl
50: X = 4-pyridyl 10: X = 4-pyridyl
51 : X = phenyl 11 : X = phenyl
52: X = 4-fluorophenyl 12: X = 4-fluorophenyl
Preparation of methyl 4-( ( 2,3-dimethyl-lH-indol-5-yl)methyl)benzoate
To a solution of N'-((2,3-dimethyl- lH-indol-5-yl)methylene)-4- methylbenzenesulfonohydrazide (3.93 g, 11.5 mmol) in 1,4-dioxane (40 mL), K2CO3 (1.7 g, 12 mmol) and (4-(methoxycarbonyl)phenyl)boronic acid (2.07 g, 11.5 mmol) were added at 80 °C. The reaction temperature was raised to 110 °C and maintained for 4 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with water and extracted with ethyl acetate. The combined organic layers were washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography, using 20-25% ethyl acetate in petroleum ether as an eluent, to afford methyl 4-((2,3 -dimethyl- lH-indol-5- yl)methyl)benzoate as a brown solid (1.5 g, 45%).
1H NMR (300 MHz, DMSO-d6): δ 10.55 (br s, 1H), 7.86 (d, = 8.7 Hz, 2H), 7.36 (d, = 8.4 Hz, 2H), 7.21 (br s, 1H), 7.12 (d, = 8.4 Hz, 1H), 6.83 (dd, = 8.4 Hz, 1.8 Hz, 1H), 4.05 (s, 2H), 3.82 (s, 3H), 2.27 (s, 3H), 2.11 (s, 3H). LCMS: m/z 294.41 [M+H]+.
Preparation of methyl 4-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoate
NaH (100 mg, 4.2 mmol) was added to dry DMF (5 niL) at 0 °C and stirred for 10 minutes. A solution of methyl 4-((2,3 -dimethyl- lH-indol-5-yl)methyl)benzoate (240 mg, 0.81 mmol) in DMF (2 mL) was added dropwise. The mixture was stirred at 0 °C for 30 minutes. To this, bromochloropropane (0.16 mL, 1.6 mmol) in DMF (2 mL) was added dropwise at 0 °C. The reaction mixture was allowed to stir at this temperature for 3 hours. After complete consumption of the starting material, the reaction mixture was quenched with saturated ammonium chloride solution, diluted with ice-cold water and then extracted with ethyl acetate. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel using 3% ethyl acetate in petroleum ether as an eluent, to afford methyl 4-((l-(3-chloropropyl)-2,3-dimethyl- lH-indol-5-yl)methyl)benzoate (130 mg, crude).
1H NMR (400 MHz, CDC13): δ 7.93 (d, = 8.4 Hz, 2H), 7.30-7.27 (m, 3H), 7.20 (d, = 8.0 Hz, 1H), 6.94 (dd, = 8.0 Hz, 1.6 Hz, 1H), 4.22 (t, = 6.8 Hz, 2H), 4.18 (s, 2H), 3.89 (s, 2H), 3.50 (t, J = 6.0 Hz, 2H), 2.35 (s, 3H), 2.22-2.16 (m, 5H). LCMS: m/z 370.1 [M+H]+.
Preparation of methyl 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)methyl)benzoate
To a stirred solution of methyl 4-((l-(3-chloropropyl)-2,3-dimethyl- lH-indol- 5-yl)methyl)benzoate (130 mg, 0.35 mmol) in acetonitrile (3 mL) at room temperature, sodium iodide (0.13 g, 0.87 mmol) and sodium carbonate (92 mg, 0.87 mmol) were added, followed by N-methylpiperazine (88 mg, 0.88 mmol). The reaction mixture was heated to 85 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford crude methyl 4-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzoate as a brown oil (180 mg, crude), which was used without further purification. LCMS: m/z [M+H]+.
Preparation of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoic acid
To a solution of methyl 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoate (200 mg, 0.46 mmol) in THF:H20:MeOH (4: 1: 1, 6 mL) was added LiOH.H20 (38 mg, 0.91 mmol) at room temperature. The reaction mixture was stirred for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, neutralised to pH~7 and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzoic acid as a brown solid (160 mg, 83%). LCMS: m/z 420.3 [M+H]+.
Preparation of Compound 49, N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-
4- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide
To a solution of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoic acid (125 mg, 0.30 mmol) and Intermediate A (84 mg, 0.38 mmol) in pyridine (3 mL), was added EDC.HC1 (280 mg, 1.5 mmol) at room temperature. The mixture was stirred at 80 °C for one hour. After complete consumption of the starting material, the reaction mass was diluted with ethyl acetate. The organic layer was washed with ammonium chloride solution, water and brine, then dried, concentrated and purified by flash column chromatography using 4% methanol in dichloromethane as an eluent to afford N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-
5- yl)methyl)benzamide (Compound 49) as an off-white solid (125 mg, 68%). 1H NMR (400 MHz, CD3OD): δ 7.86 (d, = 8.4 Hz, 2H), 7.59 (br s, 1H), 7.38 (d, = 8.0 Hz, 2H), 7.30 (s, 2H), 7.26 (br s, 1H), 7.23 (d, = 8.4 Hz, 1H), 7.05 (d, = 10.0 Hz, 2H), 6.94 (dd, = 8.4 Hz, 1.2 Hz, 1H), 4.17-4.11 (m, 4H), 3.79 (s, 6H), 2.58-2.28 (m, 16H), 2.19 (s, 3H), 1.95-1.87 (m, 2H). LCMS: m/z 621.3 [M+H]+.
Other analogues prepared by this method:
Compound 50, N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- 1- (3 -(4-methylpiperazin- 1 -yl)propyl)- lH-indol-5 -yl)methyl)benzamide (37 % ) .
1H NMR (400 MHz, CD3OD): δ 8.46 (d, = 6.0 Hz, 2H), 7.87 (d, = 8.0 Hz, 2H), 7.40-7.36 (m, 4H), 7.26-7.22 (m, 4H), 6.94 (dd, = 8.4 Hz, 1.6 Hz, 1H), 4.15- 4.11 (m, 4H), 3.76 (s, 6H), 2.58-2.29 (m, 13H), 2.27 (s, 3H), 2.19 (s, 3H), 1.95-1.87 (m, 2H). LCMS: m/z 630.4 [M-H]~.
Compound 51, N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(3- (4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide (84%).
1H NMR (400 MHz, CD3OD): δ 7.87 (d, = 8.0 Hz, 2H), 7.39 (d, = 8.4 Hz, 2H), 7.33-7.20 (m, 8H), 7.18 (s, 2H), 6.95 (dd, = 8.0 Hz, 1.2 Hz, 1H), 4.17 (t, = 6.8 Hz, 2H), 4.13 (s, 2H), 3.71 (s, 6H), 2.94-2.48 (11H), 2.40-2.35 (m, 5H), 2.20 (s, 3H), 1.98-1.89 (m, 2H). LCMS: m/z 631.4 [M+H]+.
Compound 52, 4-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (64%).
1H NMR (400 MHz, CD3OD): δ 7.86 (d, = 8.4 Hz, 2H), 7.39 (d, = 8.0 Hz, 2H), 7.27-7.22 (m, 4H), 7.18 (s, 2H), 7.04 (t, = 8.8 Hz, 2H), 6.95 (dd, = 8.4 Hz, 1.2 Hz, 1H), 4.17 (t, = 6.8 Hz, 2H), 4.13 (s, 2H), 3.72 (s, 6H), 2.94-2.47 (m, 11H), 2.41-2.35 (m, 5H), 2.20 (s, 3H), 1.92 (quintet, = 6.8 Hz, 2H). LCMS: m/z 649.3 [M+H]+.
Preparation of Compound 11, N-(2,6-dihydroxy-[l,l '-biphenyl]-4-yl)-4-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide To a stirred solution of Compound 51 (80 mg, 0.13 mmol) in dichloromethane (5 mL) at 0 °C was added boron tribromide (0.50 mL, 0.5 mmol, 1.0 M in DCM). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by prep-TLC using 5% methanol in dichloromethane as an eluent, to afford N-(2,6- dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 -(4-methylpiperazin- 1 - yl)propyl)-lH-indol-5-yl)methyl)benzamide (Compound 11) as an off white solid (18 mg, 23%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.4 Hz, 2H), 7.36-7.32 (m, 6H), 7.27-7.22 (m, 3H), 6.94 (dd, = 8.4 Hz, 1.6 Hz, 1H), 6.86 (s, 2H), 4.15 (t, = 6.8 Hz, 2H), 4.12 (s, 2H), 2.62-2.39 (m, 8H), 2.35-2.30 (m, 8H), 2.19 (s, 3H), 1.90 (quintet, = 7.2 Hz, 2H). LCMS: m/z 603.3 [M+H]+.
Other analogues prepared by this method:
Compound 9, N-(3,5-dihydroxy-4-(lH-imidazol- l-yl)phenyl)-4-((2,3- dimethyl-1 -(3 -(4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)methyl)benzamide (4%).
1H NMR (400 MHz, CD3OD): δ 7.80 (d, = 8.4 Hz, 2H), 7.65 (br s, 1H), 7.36 (d, = 8.0 Hz, 2H), 7.25-7.22 (m, 2H), 7.13 (br s, 1H), 7.05 (br s, 1H), 6.96 (s, 2H), 6.93 (br d, = 10.0 Hz, 1H), 4.15-4.10 (m, 4H), 2.69-2.40 (m, 8H), 2.35 (s, 3H), 2.33-2.29 (m, 5H), 2.19 (s, 3H), 1.95-1.85 (m, 2H). LCMS: m/z 593.2 [M+H]+.
Compound 10, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (3 -(4-methylpiperazin- 1 -yl)propyl)- lH-indol-5-yl)methyl)benzamide ( 1 %) .
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 6.0 Hz, 2H), 7.80 (d, = 8.0 Hz, 2H), 7.53 (d, = 6.0 Hz, 2H), 7.36 (d, = 8.4 Hz, 2H), 7.25 (br s, 1H), 7.23 (d, = 8.4 Hz, 1H), 6.93 (br d, = 8.4 Hz, 1H), 6.90 (s, 2H), 4.20-4.11 (m, 4H), 2.85-2.28 (m, 16H), 2.19 (s, 3H), 1.97-1.85 (m, 2H). LCMS: m/z 604.3 [M+H]+. Compound 12, 4-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4-yl)benzamide (23%).
1H NMR (400 MHz, DMSO-d6): δ 9.96 (br s, 1H), 9.20 (br s, 2H), 7.82 (d, = 8.0 Hz, 2H), 7.39-7.31 (m, 4H), 7.28-7.25 (m, 2H), 7.12 (t, = 8.8 Hz, 2H), 6.93- 6.90 (m, 3H), 4.08-4.04 (m, 4H), 2.38-2.14 (m, 19H), 1.78-1.71 (m, 2H). LCMS: m/z 621.3 [M+H]+.
Scheme 11. Preparation of Compounds 14-16 and 53-56
Figure imgf000071_0001
53: X = 1-imidazolyl 14: X = 4-pyridyl
54: X = 4-pyridyl 15: X = phenyl
55: X = phenyl 16: X = 4-fluorophenyl
56: X = 4-fluorophenyl
Preparation of methyl 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl )propyl)-lH-indol-5-yl )methyl )benzoate
Tosyl hydrazide (2.3 g, 12 mmol) was added to a stirred solution of 2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole-5-carbaldehyde (2.7 g, 8.2 mmol) in dry 1,4-dioxane (60 mL) at room temperature, followed by the addition of glacial acetic acid (0.7 mL, cat.). The mixture was stirred at room temperature for 3 hours. To this solution was added K2CO3 (4.5 g, 33 mmol) and (4-(methoxycarbonyl)phenyl)boronic acid (2.2 g, 12 mmol). The reaction temperature was raised to 110 °C and maintained for 5 hours. After complete consumption of the starting material, the reaction mass was concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 10% methanol in dichloromethane as an eluent, to obtain methyl 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)methyl)benzoate as a yellow solid (1.4 g, 38%). LCMS: m/z 448.5 [M+H]+.
Preparation of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl )propyl)-lH-indol-5-yl )methyl )benzoic acid
To a solution of methyl 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoate (1.4 g, 3.1 mmol) in THF:H20:MeOH (4: 1: 1, 12 niL) was added 2 N NaOH solution (4 mL) at 0 °C. The reaction mixture was stirred at room temperature for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, neutralised to pH~7 and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 10% methanol in dichloromethane as an eluent, to afford 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoic acid as an off white solid (1.0 g, 74%). LCMS: m/z 434.42 [M+H]+.
Preparation of Compound 53, N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)- 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide
To a solution of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)benzoic acid (250 mg, 0.58 mmol) and Intermediate A (165 mg, 0.75 mmol) in pyridine (5 mL), was added EDC.HC1 (660 mg, 3.4 mmol). The mixture was stirred at room temperature for 10 minutes, then heated to 80 °C and stirred for 3 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with water and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, then dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 5-10% methanol in dichloromethane as an eluent, to afford N- (4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (Compound 53) as an off-white solid (200 mg, 55%).
1H NMR (400 MHz, DMSO-d6): δ 10.29 (br s, IH), 7.90 (d, = 8.4 Hz, 2H), 7.54 (br s, IH), 7.43-7.38 (m, 4H), 7.28 (br s, IH), 7.23 (d, = 8.4 Hz, IH), 7.09 (br s, IH), 6.98 (br s, IH), 6.92 (dd, = 8.4 Hz, 1.6 Hz, IH), 4.14 (dd, = 15.2 Hz, 4.8 Hz, IH), 4.08 (s, 2H), 3.82-3.76 (m, IH), 3.72 (s, 6H), 2.49-2.11 (m, 20H), 0.73 (d, = 6.4 Hz, 3H). LCMS: m/z 633.61 [M+H]+.
Other analogues prepared by this method:
Compound 54, N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl- 1- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (19%).
1H NMR (300 MHz, DMSO-d6): δ 10.34 (br s, IH), 8.53 (d, = 5.4 Hz, 2H), 7.90 (d, = 8.4 Hz, 2H), 7.42 (d, = 8.1 Hz, 2H), 7.36 (s, 2H), 7.28-7.22 (m, 4H), 6.92 (br d, = 7.5 Hz, IH), 4.14 (dd, = 14.7 Hz, 6.4 Hz, IH), 4.08 (s, 2H), 3.79 (dd, = 14.7 Hz, 8.4 Hz, IH), 3.69 (s, 6H), 2.49-2.14 (m, 20H), 0.73 (d, = 6.3 Hz, 3H). LCMS: m/z 644.76 [M+H]+.
Compound 55, N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2- methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (21%).
1H NMR (300 MHz, DMSO-d6): δ 10.19 (br s, IH), 7.89 (d, = 8.1 Hz, 2H), 7.41 (d, = 8.1 Hz, 2H), 7.39-7.19 (m, 9H), 6.93 (br d, = 8.7 Hz, IH), 4.14 (dd, = 14.4 Hz, 4.2 Hz, IH), 4.08 (s, 2H), 3.79 (dd, = 14.7 Hz, 8.7 Hz, IH), 3.65 (s, 6H), 2.49-2.12 (m, 20H), 0.73 (d, = 6.3 Hz, 3H). LCMS: m/z 645.64 [M+H]+.
Compound 56, 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4- yl)benzamide (47%). 1H NMR (300 MHz, DMSO-d6): δ 10.19 (br s, 1H), 7.89 (d, = 8.4 Hz, 2H), 7.41 (d, = 8.7 Hz, 2H), 7.32 (s, 2H), 7.30-7.22 (m, 6H), 7.16 (t, = 9.3 Hz, 2H), 6.92 (br d, J = 8.4 Hz, 1H), 4.14 (dd, = 15.0 Hz, 4.8 Hz, 1H), 4.08 (s, 2H), 3.79 (dd, = 14.7 Hz, 8.7 Hz, 1H), 3.66 (s, 6H), 2.49-2.12 (m, 20H), 0.73 (d, = 6.3 Hz, 3H). LCMS: m/z 663.60 [M+H]+.
Preparation of Compound 14, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide
To a stirred solution of Compound 54 (50 mg, 0.078 mmol) in dichloromethane (3 mL) at 0 °C was added boron tribromide (2.0 mL, 2.0 mmol,
1.0 M in DCM). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, then dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by prep-TLC, using 5% methanol in dichloromethane as an eluent, to afford N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)methyl)benzamide (Compound 14) as an off white solid (17 mg, 35%).
1H NMR (300 MHz, DMSO-d6): δ 10.03 (br s, 1H), 9.48 (br s, 2H), 8.48 (d, = 6.0 Hz, 2H), 7.83 (d, = 8.1 Hz, 2H), 7.40-7.34 (m, 4H), 7.27 (br s, 1H), 7.23 (d, = 8.1 Hz, 1H), 6.98 (s, 2H), 6.91 (br d, = 8.1 Hz, 1H), 4.14 (dd, = 14.4 Hz, 4.8 Hz, 1H), 4.07 (s, 2H), 3.79 (dd, = 13.8 Hz, 8.7 Hz, 1H), 2.49-2.05 (m, 20H), 0.73 (d, = 6.3 Hz, 3H). LCMS: m/z 618.46 [M+H]+.
Other analogues prepared by this method:
Compound 15, N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(2- methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)methyl)benzamide (43%).
1H NMR (300 MHz, DMSO-d6): δ 9.96 (br s, 1H), 9.13 (br s, 2H), 7.83 (d, =
8.1 Hz, 2H), 7.38 (d, = 8.1 Hz, 2H), 7.33-7.20 (m, 7H), 6.94-6.89 (m, 3H), 4.18- 4.04 (m, 3H), 3.83-3.74 (m, 1H), 2.42-2.07 (m, 20H), 0.73 (d, = 6.0 Hz, 3H). LCMS: m/z 617.44 [M+H]+.
Compound 16, 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l- yl)propyl)- lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4- yl)benzamide (17%).
1H NMR (300 MHz, DMSO-d6): δ 9.96 (br s, 1H), 9.21 (br s, 2H), 7.82 (d, = 8.1 Hz, 2H), 7.38 (d, = 8.4 Hz, 2H), 7.33 (dd, = 8.7 Hz, 5.7 Hz, 2H), 7.27 (br s, 1H), 7.23 (d, = 8.4 Hz, 1H), 7.12 (t, / = 8.7 Hz, 2H), 6.94-6.89 (m, 3H), 4.14 (dd, = 14.7 Hz, 4.5 Hz, 1H), 4.06 (s, 2H), 3.78 (s, / = 14.7 Hz, 9.0 Hz, 1H), 2.45-2.07 (m, 20H), 0.72 (d, J = 6.0 Hz, 3H). LCMS: m/z 636.40 [M+H]+.
Scheme 12. Preparation of Compounds 17-20.
Figure imgf000075_0001
Preparation of 5-methoxy-2,3-dimethyl-lH-indole
2-Butanone (19.3 mL, 0.215 mol) was added to a stirred solution of 4-methoxyhydrazine hydrochloride (25.0 g, 0.143 mol) in acetic acid (250 mL), then heated at 130 °C for 3 hours. After complete consumption of the starting material, the reaction mixture was quenched with water and stirred for 30 minutes at room temperature. The resulting precipitate was collected by filtration and dried for one hour to afford 5-methoxy-2,3-dimethyl-lH-indole as a brown solid (22 g, 88%). LCMS: m/z 176.1 [M+H]+.
Preparation of2,3-dimethyl-lH-indol-5-ol
BBr3 (142 mL, 1.0 M in DCM) was added to a stirred solution of 5-methoxy- 2,3-dimethyl-lH-indole (10.0 g, 57.1 mmol) in dichloromethane (200 mL) at 0 °C. The temperature was maintained at 0-5 °C for 3 hours. After complete consumption of the starting material, the reaction mixture was basified with saturated NaHC03 then extracted with dichloromethane. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography, using 10% ethyl acetate in petroleum ether as an eluent, to afford 2,3-dimethyl-lH- indol-5-ol as a brown solid (8.2 g, 89%).
1H NMR (400 MHz, DMSO-d6): δ 10.26 (s, 1H), 8.44 (s, 1H), 6.98 (d, =
8.4 Hz, 1H), 6.64 (d, = 2.4 Hz, 1H), 6.47 (dd, = 8.4 Hz, 2.4 Hz, 1H), 2.24 (s, 3H),
2.05 (s, 3H). LCMS: m/z 162.2 [M+H]+.
Preparation oftert-butyl 5-hydroxy-2,3-dimethyl-lH-indole-l -carboxylate
To a stirred solution of 2,3-dimethyl-lH-indol-5-ol (8.2 g, 51 mmol), in acetonitrile (80 mL) was added Boc-anhydride (32.9 g, 151 mmol) and DMAP (0.61 g, 5.0 mmol) at room temperature. The reaction mass was stirred at room temperature for 16 hours. After complete consumption of the starting material, acetonitrile was evaporated under reduced pressure. The residue was re-dissolved in methanol (200 mL), K2C03 (20.7 g, 150 mmol) was added and the resulting mixture was stirred at room temperature for 4 hours. After completion of the reaction, the reaction mass was extracted with ethyl acetate. The organic layer was washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 13% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-hydroxy-2,3-dimethyl- lH-indole- 1-carboxylate as a yellow solid (11.2 g, 84%).
1H NMR (400 MHz, CDC13): δ 7.93 (d, = 8.8 Hz, 1H), 6.83 (d, = 2.4 Hz, 1H), 6.73 (dd, = 8.8 Hz, 2.8 Hz, 1H), 4.77 (s, 1H), 2.50 (s, 3H), 2.12 (s, 3H), 1.66 (br s, 9H). LCMS: m/z 262.1 [M+H]+.
Preparation of tert-butyl 5-(3-(methoxycarbonyl)phenoxy)-2,3-dimethyl-lH- indole-l -carboxylate
To a stirred solution of tert-butyX 5-hydroxy-2,3-dimethyl- lH-indole- l- carboxylate (5.0 g, 19 mmol) in dichloromethane (150 mL) was added 3-(methoxycarbonyl)phenyl)boronic acid (10.3 g, 57.2 mmol). Cu(OAc)2 (8.7 g, 48 mmol) was then added, followed by triethylamine (13.3 mL, 95 mmol) and the system was purged with oxygen gas for 4 hours. The whole reaction mass was stirred under an oxygen atmosphere for a further 16 hours. After complete consumption of the starting material, the reaction mass was filtered through a bed of Celite. The filtrate was diluted with water and extracted with dichloromethane. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 3% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-(3-(methoxycarbonyl)phenoxy)-2,3-dimethyl- lH-indole-1 -carboxylate as a brown liquid (3.5 g, 46%).
1H NMR (400 MHz, CDC13): δ 8.08 (d, = 8.8 Hz, 1H), 7.22 (dt, = 7.6 Hz, 1.2 Hz, 1H), 7.60 (dd, 7 = 2.4 Hz, 1.6 Hz, 1H), 7.36 (t, J = 8.0 Hz, 1H), 7.19-7.15 (m, 1H), 7.07 (d, = 2.4 Hz, 1H), 6.93 (dd, = 8.8 Hz, 1.6 Hz, 1H), 3.87 (s, 3H), 2.53 (s, 3H), 2.13 (s, 3H), 1.68 (s, 9H). LCMS: m/z 396.1 [M+H]+.
Preparation of methyl 3-( ( 2,3-dimethyl-lH-indol-5-yl)oxy)benzoate
To a stirred solution of tert-butyX 5-(3-(methoxycarbonyl)phenoxy)-2,3- dimethyl-lH- indole- 1 -carboxylate (3.5 g, 20.8 mmol) in dichloromethane (10 mL) at 0 °C, was added dropwise 1,4-dioxane hydrochloride (25 mL, 4.0 M in dichloromethane). The mixture was allowed to warm to room temperature and stirred for 20 hours. After complete consumption of the starting material, the reaction mixture was concentrated in vacuo, basified to pH 10 with aqueous NaHC03 and extracted with ethyl acetate. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford methyl 3-((2,3-dimethyl-lH-indol- 5-yl)oxy)benzoate as a brown liquid (2.5 g, 96%).
1H NMR (400 MHz, CDC13): δ 7.75 (br s, 1H), 7.68 (d, = 8.0 Hz, 1H), 7.60 (t, = 2.4 Hz, 1H), 7.33 (t, = 8.0 Hz, 1H), 7.23 (d, = 8.4 Hz, 1H), 7.17-7.13 (m, 2H), 6.84 (dd, = 8.4 Hz, 2.4 Hz, 1H), 3.86 (s, 3H), 2.38 (s, 3H), 2.17 (s, 3H). LCMS: m/z 296.1 [M+H]+.
Preparation of methyl 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoate
To a stirred suspension of NaH (1.17 g, 49 mmol) in dry DMF (lOmL) at 0 °C was added dropwise a solution of methyl 3-((2,3-dimethyl-lH-indol-5- yl)oxy)benzoate (2.4 g, 8.1 mmol) in DMF (10 mL). The mixture was allowed to warm to room temperature for 30 minutes. To this, bromochloropropane (1.6 mL, 16 mmol) was added dropwise at 0 °C. The mixture was stirred for 2 hours. After complete consumption of the starting material, ice-cold water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with chilled ammonium chloride solution and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude product was purified by flash column chromatography, using 4% ethyl acetate in petroleum ether as an eluent, to afford methyl 3-((l-(3-chloropropyl)-2,3-dimethyl- lH-indol-5-yl)oxy)benzoate as a yellow liquid (600 mg, 20%).
1H NMR (400 MHz, CDC13): δ 7.69 (dt, = 7.6 Hz, 1.2 Hz, 1H), 7.60 (t, = 2.0 Hz, 1H), 7.33 (t, J = 8.0 Hz, 1H), 7.26-7.18 (m, 1H), 7.17-7.14 (m, 2H), 6.87 (dd, = 8.8 Hz, 2.4 Hz, 1H), 4.26 (t, = 6.8 Hz, 2H), 3.87 (s, 3H), 3.55 (t, = 6.0 Hz, 2H), 2.38 (s, 3H), 2.25-2.19 (m, 5H). LCMS: m/z 372.1 [M+H]+. Preparation of methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)oxy)benzoate
To a stirred solution of methyl 3-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol- 5-yl)oxy)benzoate (600 mg, 1.6 mmol) in acetonitrile (10 mL), were added sodium iodide (600 mg, 4.0 mmol) and sodium carbonate (420 mg, 3.9 mmol) followed by N-methylpiperazine (400 mg, 4.0 mmol) at room temperature. The reaction mixture was heated to 85 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography, using 5% methanol in dichloromethane as a eluent, to afford methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzoate (380 mg, 44%).
1H NMR (400 MHz, CDC13): δ 7.68 (dt, = 7.6 Hz, 1.2 Hz, 1H), 7.59 (t, = 2.4 Hz, 1H), 7.33 (t, = 8.0 Hz, 1H), 7.26-7.18 (m, 1H), 7.17-7.14 (m, 2H), 6.85 (dd, = 8.8 Hz, 2.4 Hz, 1H), 4.13 (t, = 7.2 Hz, 2H), 3.86 (s, 3H), 2.75-2.39 (m, 8H), 2.38-2.20 (m, 8H), 2.17 (s, 3H), 1.95-1.88 (m, 2H). LCMS: m/z 436.3 [M+H]+.
Preparation of 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)oxy)benzoic acid
To a solution of methyl 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoate (380 mg, 0.87 mmol) in THF:H20:MeOH (4: 1: 1, 10 mL) was added LiOH.H20 (360 mg, 8.6 mmol) at room temperature. The reaction mixture was stirred for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, cooled to 0 °C, acidified to pH 2 with 1 N HC1 and extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by trituration with n-pentane to afford 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzoic acid as a brown solid (320 mg, 87%). LCMS: m/z 422.1 [M+H]+. Preparation of N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a solution of 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)oxy)benzoic acid (97 mg, 0.23 mmol) and Intermediate B (69 mg, 0.30 mmol) in pyridine (1 mL), was added EDC.HCl (265 mg, 1.4 mmol). The mixture was stirred at room temperature for 10 minutes, then heated to 80 °C for one hour. After complete consumption of the starting material, the reaction mass was concentrated, cooled to 0 °C, diluted with water and extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using ethyl acetate and 5% methanol in dichloromethane as an eluent, to afford N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl- 1-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide as a brown solid (50 mg, 34%).
1H NMR (400 MHz, DMSO-d6): δ 10.32 (br s, 1H), 8.53 (dd, = 8.4 Hz, 1.6 Hz, 2H), 7.67 (d, = 8.0 Hz, 1H), 7.51-7.42 (m, 3H), 7.34 (s, 2H), 7.25 (dd, = 8.4 Hz, 1.6 Hz, 2H), 7.15-7.12 (m, 2H), 6.83 (dd, = 8.8 Hz, 2.4 Hz, 1H), 4.14 (t, = 6.8 Hz, 2H), 3.68 (s, 6H), 2.46-2.15 (m, 19H), 1.80 (quintet, = 6.8 Hz, 2H). LCMS: m/z 634.3 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (41%). LCMS: m/z 623.3 [M+H]+.
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (66%).
1H NMR (400 MHz, DMSO-d6): δ 10.27 (br s, 1H), 7.67 (d, = 8.0 Hz, 1H), 7.51-7.43 (m, 3H), 7.35-7.30 (m, 4H), 7.26-7.12 (m, 5H), 6.83 (dd, = 8.8 Hz, 1.6 Hz, 1H), 4.14 (br s, 2H), 3.64 (s, 6H), 2.46-2.14 (m, 19H), 1.81 (br s, 2H). 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)-N- (4'-fluoro-2,6-dimethoxy- [1,1 '-biphenyl] -4-yl)benzamide (35%).
1H NMR (400 MHz, DMSO-d6): δ 10.27 (br s, 1H), 7.68 (d, = 7.6 Hz, 1H), 7.51-7.43 (m, 3H), 7.25-7.21 (m, 2H), 7.17-7.12 (m, 4H), 6.84 (dd, = 8.4 Hz, 2.4 Hz, 1H), 4.14 (t, = 6.8 Hz, 2H), 3.65 (s, 6H), 2.81-2.24 (m, 16H), 2.15 (s, 3H), 1.88-1.77 (m, 2H). LCMS: m/z 651.3 [M+H]+.
Preparation of Compound 19, N-(2,6-dihydroxy- [ 1 ,1 '-biphenyl] -4-yl)-3 '-((2,3 '- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a stirred solution of N-(2,6-dimethoxy-[l, -biphenyl]-4-yl)-3-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
(110 mg, 0.17 mmol) in dichloromethane (2 mL) at 0 °C was added boron tribromide (0.50 mL, 5.3 mmol). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by prep-TLC, using 2.5% methanol in dichloromethane as an eluent, to afford N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (Compound 19) as an off-white solid (20 mg, 19%).
1H NMR (400 MHz, CD3OD): δ 7.54 (d, = 7.6 Hz, 1H), 7.45-7.34 (m, 7H), 7.28-7.23 (m, 1H), 7.16-7.12 (m, 2H), 6.85 (dd, = 8.8 Hz, 2.4 Hz, 1H), 6.81 (s, 2H), 4.24 (t, = 6.8 Hz, 2H), 3.21-2.32 (m, 16H), 2.19 (s, 3H), 1.97 (quintet, = 6.8 Hz, 2H). LCMS: m/z 605.3 [M+H]+.
Other analogues prepared by this method:
Compound 17, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-3-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (5%). 1H NMR (400 MHz, CD3OD): δ 7.64 (br s, IH), 7.54 (br d, J = 8.0 Hz, IH), 7.45-7.39 (m, 2H), 7.36 (d, J = 8.4 Hz, IH), 7.14-7.10 (m, 3H), 7.05 (br s, IH), 6.87 (s, 2H), 6.84 (dd, = 8.4 Hz, 2.0 Hz, IH), 4.19 (t, = 6.8 Hz, 2H), 2.75-2.41 (m, 8H), 2.38 (s, 3H), 2.34-2.30 (m, 5H), 2.19 (s, 3H), 1.97-1.86 (m, 2H). LCMS : m/z 595.3 [M+H]+.
Compound 18, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (3 -(4-methylpiperazin- 1 -yl)propyl)- lH-indol-5 -yl)oxy)benzamide (21%).
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 5.6 Hz, 2H), 7.56-7.50 (m, 3H), 7.44-7.39 (m, 2H), 7.36 (d, / = 8.8 Hz, IH), 7.13-7.10 (m, 2H), 6.87 (s, 2H), 6.83 (dd, = 8.8 Hz, 2.4 Hz, IH), 4.20 (t, = 6.8 Hz, 2H), 2.74-2.41 (m, 8H), 2.38 (s, 3H), 2.34 (t, = 7.2 Hz, 2H), 2.31 (s, 3H), 2.19 (s, 3H), 1.94 (quintet, = 7.2 Hz, 2H). LCMS: m/z 606.3 [M+H]+.
Compound 20, 3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)oxy)-N-(4'-fluoro-2,6-dihydroxy-[l, l'-biphenyl]-4-yl)benzamide (12%).
1H NMR (400 MHz, CD3OD): δ 7.53 (d, = 8.0 Hz, IH), 7.45-7.33 (m, 5H), 7.13-7.09 (m, 2H), 7.06 (t, J = 8.8 Hz, 2H), 6.85-6.82 (m, 3H), 4.20 (t, J = 6.8 Hz, 2H), 2.98-2.40 (m, 8H), 2.38 (s, 3H), 2.36-2.31 (m, 5H), 2.19 (s, 3H), 1.94 (quintet, J = 6.8 Hz, 2H). LCMS: m/z 623.3 [M+H]+.
Scheme 13. Preparation of Compounds 22-24
Figure imgf000083_0001
Preparation of 5-methoxy-2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indole
To a stirred suspension of NaH (6.1 g, 265 mmol) in DMF (50 niL) at 0 °C was added dropwise a solution of 5-methoxy-2,3-dimethyl-lH-indole (15 g, 86 mmol) in DMF (50 mL) at 0 °C. The mixture was stirred for 30 minutes. To this, l-bromo-3- chloro-2-methylpropane (20 mL, 171 mmol) was added dropwise at 0 °C. The mixture was allowed to warm to room temperature and stirred for 16 hours. After complete consumption of the starting material, ice-cold water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with chilled ammonium chloride solution, then brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compounds was purified by column chromatography on 100-200 mesh silica gel using petroleum ether as an eluent, to afford 5-bromo-l-(3-chloro-2- methylpropyl)-2,3-dimethyl-lH-indole as a yellow liquid (6 g, crude).
To a stirred solution of the crude intermediate (6 g) in acetonitrile (100 mL), sodium iodide (8.45 g, 56.4 mmol), sodium carbonate (5.97 g, 56.3 mmol) and then N-methylpiperazine (5.65 g, 56.4 mmol) were added at room temperature. The reaction mixture was heated to 85 °C for 12 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as an eluent, to afford 5 -methoxy-2,3 -dimethyl- 1- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indole as a yellow liquid (1.75 g, 6%). LCMS: m/z 330.30 [M+H]+.
Preparation of 2, 3 -dimethyl- 1 -(2-methyl-3-(4-methylpiperazin-l -yl)propyl)- lH-indol-5-ol
To a stirred solution of 5-methoxy-2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indole (4.1 g, 12 mmol) in dichloromethane (100 mL) at -78 °C was added boron tribromide (18.6 mL, 18.6 mmol, 1 M in DCM). The mixture was stirred at this temperature for 2 hours, then slowly warmed to room temperature and stirred for another 2 hours. The reaction mixture was quenched with ice-cold saturated NaHC03 solution, then extracted with ethyl acetate. The organic layers were washed with chilled brine solution, and dried over Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 6% methanol in dichloromethane as an eluent, to afford 2,3-dimethyl-l-(2-methyl-3- (4-methylpiperazin-l-yl)propyl)-lH-indol-5-ol as a brown solid (2.1 g, 53%). LCMS: m/z 316.29 [M+H]+.
Preparation of 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzonitrile
To a stirred solution of NaH (130 mg, 5.4 mmol) in DMF (5 mL) was added a solution of 2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- ol (1.00 g, 3.17 mmol) in DMF (5 mL) dropwise at 0 °C, under an atmosphere of N2. 3-Chlorobenzonitrile (77 mg, 0.48 mmol) was added at 0 °C. The mixture was heated to 100 °C and stirred at this temperature for 12 hours. After complete consumption of the starting material, the reaction mass was cooled to 0 °C, quenched with saturated ammonium chloride solution and extracted with ethyl acetate. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 3% methanol in dichloromethane as an eluent, to afford 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzonitrile as a brown gummy solid (850 mg, 65%). LCMS: m/z 417.33 [M+H]+.
Preparation of 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoic acid
To a solution of 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzonitrile (900 mg, 2.1 mmol) in ethanol (15 mL) was added 8 N ΚΟΗ solution (15 mL) at 0 °C. The reaction mixture was heated to 100 °C for 12 hours. After complete consumption of the starting material, the reaction mass was concentrated and acidified with 6 N HC1. The residue was extracted with 30% isopropyl alcohol in chloroform, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)oxy)benzoic acid as an off white solid (800 mg, 85%). LCMS: m/z 436.38 [M+H]+.
Preparation of N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a solution of 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoic acid (200 mg, 0.45 mmol) and Intermediate B (136 mg, 0.59 mmol) in pyridine (1 mL), was added EDC.HC1 (527 mg, 2.75 mmol). The mixture was stirred at room temperature for 10 minutes, then heated to 100 °C for 2 hours. After complete consumption of the starting material, the reaction mass was concentrated, diluted with aqueous ammonium chloride and extracted with ethyl acetate. The organic layer was dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 3% methanol in dichloromethane as an eluent, to afford N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide as a brown solid (200 mg, 68%). LCMS: m/z 647.33 [Μ-Η]".
Other analogues prepared by this method:
N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (68%). LCMS: m/z 647.47 [M+H]+.
3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)-N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (66%). LCMS: m/z 665.41 [M+H]+.
Preparation of Compound 23, N-(2,6-dihydroxy-[l,l '-biphenyl]-4-yl)-3-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lll-indol-5-yl)oxy)benzamide
To a stirred solution of N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-3-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide (200 mg, 0.31 mmol) in dichloromethane (6 mL) at 0 °C was added boron tribromide (2.0 mL, 2.0 mmol, 1.0 M in DCM). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by prep-TLC using 5% methanol in dichloromethane as an eluent to afford N-(2,6-dihydroxy-[l,l'-biphenyl]-4-yl)-3- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide (Compound 23) as a pale brown solid (40 mg, 21%).
1H NMR (400 MHz, DMSO-d6): δ 10.06 (br s, 1H), 9.14 (br s, 2H), 7.60 (d, = 8.0 Hz, 1H), 7.45-7.43 (m, 2H), 7.39 (d, = 8.4 Hz, 1H), 7.30-7.28 (m, 4H), 7.25- 7.17 (m, 1H), 7.12 (d, = 2.4 Hz, 1H), 7.07 (dd, / = 8.0 Hz, 2.0 Hz, 1H), 6.92 (s, 2H), 6.82 (dd, = 8.8 Hz, 2.0 Hz, 1H), 4.18 (dd, = 14.8 Hz, 4.8 Hz, 1H), 3.86 (dd, = 14.4 Hz, 8.8 Hz, 1H), 2.52-2.08 (m, 20H), 0.78 (d, = 6.4 Hz, 3H). LCMS: m/z 619.45 [M+H]+. Other analogues prepared by this method:
Compound 22, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (7%)
1H NMR (400 MHz, DMSO-d6): δ 10.12 (br s, 1H), 9.55 (br s, 2H), 8.49 (d, = 6.0 Hz, 1H), 7.60 (d, = 8.0 Hz, 1H), 7.47-7.34 (m, 5H), 7.12 (d, = 2.4 Hz, 1H), 7.08 (dd, = 8.4 Hz, 2.0 Hz, 1H), 6.97 (s, 2H), 6.82 (dd, = 8.8 Hz, 2.4 Hz, 1H), 4.18 (dd, = 14.8 Hz, 4.4 Hz, 1H), 3.95 (dd, = 14.4 Hz, 8.4 Hz, 1H), 2.52-2.14 (m, 20H), 0.78 (d, J = 6.4 Hz, 3H). LCMS: m/z 620.39 [M+H]+.
Compound 24, 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l- yl)propyl)- lH-indol-5-yl)oxy)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4- yl)benzamide (31%)
1H NMR (400 MHz, DMSO-d6): δ 10.07 (br s, 1H), 9.22 (br s, 2H), 7.60 (d, = 8.0 Hz, 1H), 7.46-7.42 (m, 2H), 7.38 (d, = 8.8 Hz, 1H), 7.34-7.30 (m, 2H), 7.15- 7.10 (m, 3H), 7.07 (dd, = 8.4 Hz, 2.4 Hz, 1H), 6.93 (s, 2H), 6.82 (dd, = 8.8 Hz, 2.0 Hz, 1H), 4.19 (dd, = 14.4 Hz, 4.4 Hz, 1H), 3.95 (dd, = 14.4 Hz, 8.4 Hz, 1H), 2.52-2.09 (m, 20H), 0.78 (d, = 5.6 Hz, 3H). LCMS: m/z 637.46 [M+H]+.
Scheme 14. Preparation of Compounds 26-28
Figure imgf000087_0001
Figure imgf000087_0002
Preparation of tert-butyl 5-(4-(methoxycarbonyl)phenoxy)-2,3-dimethyl-lH- indole-l -carboxylate
To a stirred solution of tert-butyX 5-hydroxy-2,3-dimethyl- lH-indole- l- carboxylate (6.0 g, 23 mmol) in dichloromethane (300 mL) was added 4-(methoxycarbonyl)phenyl)boronic acid (12.4 g, 68.9 mmol). Cu(OAc)2 (10.4 g, 57.3 mmol) was then added, followed by triethylamine (9.6 mL, 69 mmol) and the system was purged with oxygen gas for 4 hours. The whole reaction mass was stirred under an oxygen atmosphere for a total of 16 hours. After complete consumption of the starting material, the reaction mass was filtered through a bed of Celite. The filtrate was diluted with water and extracted with dichloromethane. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 10% ethyl acetate in petroleum ether as an eluent, to afford tert-butyX 5-(4-(methoxycarbonyl)phenoxy)-2,3-dimethyl- lH-indole-1 -carboxylate as an off white solid (6.0 g, 66%).
1H NMR (400 MHz, CDC13): δ 8.10 (d, = 8.8 Hz, 1H), 7.97 (d, = 8.8 Hz, 2H), 7.11 (d, J = 2.0 Hz, 1H), 6.97-6.93 (m, 3H), 3.89 (s, 3H), 2.53 (s, 3H), 2.14 (s, 3H), 1.68 (s, 9H). LCMS: m/z 396.1 [M+H]+.
Preparation of methyl 4-( ( 2,3-dimethyl-lH-indol-5-yl)oxy)benzoate
To a stirred solution of tert-butyX 5-(4-(methoxycarbonyl)phenoxy)-2,3- dimethyl-lH- indole- 1 -carboxylate (6.0 g, 15 mmol) in dichloromethane (100 mL) at 0 °C was added dropwise 1,4-dioxane hydrochloride (25 mL, 4.0 M in DCM). The mixture was allowed to warm to room temperature and stirred for 20 hours. The reaction mixture was concentrated in vacuo, basified to pH 10 with aqueous NaHC03 solution and extracted with ethyl acetate. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford methyl 4-((2,3- dimethyl-lH-indol-5-yl)oxy)benzoate as an off white solid (4.2 g, 94%). LCMS: m/z 296.1 [M+H]+. Preparation of methyl 4-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoate
To a stirred suspension of NaH (1.7 g, 71 mmol) in DMF (20 mL) was added dropwise a solution of methyl 4-((2,3-dimethyl-lH-indol-5-yl)oxy)benzoate (4.2 g, 14 mmol) in DMF (10 mL) at 0 °C. The mixture was stirred at room temperature for 30 minutes, then cooled to 0 °C. To this mixture, bromochloropropane (3.2 mL, 32 mmol) was added dropwise. The mixture was stirred for 2 hours. After complete consumption of the starting material, ice-cold water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with chilled ammonium chloride solution, then brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography, using 4% ethyl acetate in petroleum ether as an eluent, to afford methyl 4-((l-(3-chloropropyl)-2,3-dimethyl- lH-indol-5-yl)oxy)benzoate as a black solid (2.5 g, 48%). LCMS: m/z 372.1 [M+H]+.
Preparation of methyl 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)oxy)benzoate
To a stirred solution of methyl 4-((l-(3-chloropropyl)-2,3-dimethyl-lH-indol- 5-yl)oxy)benzoate (2.0 g, 5.4 mmol) in acetonitrile (20 mL), were added sodium iodide (2.1 g, 14 mmol) and sodium carbonate (1.5 g, 14 mmol), followed by N-methylpiperazine (1.6 mL, 14 mmol) at room temperature. The reaction mixture was heated to 85 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography using 5% methanol in dichloromethane as the eluent to afford methyl 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzoate as a yellow liquid (1.1 g, 47%). LCMS: m/z 436.3 [M+H]+.
Preparation of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)oxy)benzoic acid To a stirred solution of methyl 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoate (1.1 g, 2.5 mmol) in THF:MeOH:H20 (4: 1: 1, 16 mL) was added aqueous 2 N NaOH solution (8 mL) at 0 °C. The reaction mixture was allowed to warm to room temperature and was stirred for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated in vacuuo, cooled to 0 °C, acidified with 1 N HC1 up to pH 2 and extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 4-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzoic acid as an off white solid (0.80 g, 75%). LCMS: m/z 422.1 [M+H]+.
Preparation of N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a solution of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)oxy)benzoic acid (150 mg, 0.36 mmol) and Intermediate B (110 mg, 0.48 mmol) in pyridine (10 mL), was added EDC.HC1 (410 mg, 21 mmol). The mixture was stirred at room temperature for 10 minutes, then heated to 80 °C for one hour. After complete consumption of the starting material, the reaction mixture was quenched with aqueous ammonium chloride and extracted with ethyl acetate. The organic layer was washed with water and brine, then dried over anhydrous Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as an eluent, to afford N-(3,5-dimethoxy-4-(pyridin- 4-yl)phenyl)-4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide as a brown solid (100 mg, 44%).
1H NMR (400 MHz, CDC13): δ 8.60 (d, = 4.4 Hz, 2H), 7.84 (d, = 8.4 Hz, 2H), 7.30-7.21 (m, 4H), 7.06 (br s, 2H), 7.01 (d, / = 8.4 Hz, 2H), 6.88 (br d, / = 8.4 Hz, 1H), 4.16 (t, = 6.4 Hz, 2H), 3.78 (s, 6H), 2.83-2.27 (m, 16H), 2.21 (s, 3H), 1.98-1.84 (m, 2H). LCMS: m/z 634.37 [M+H]+.
Other analogues prepared by this method: N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (44%). LCMS: m/z 633.48 [M+H]+.
4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)-N- (4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (39%). LCMS: m/z 651.3 [M+H]+.
Preparation of Compound 26, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a stirred solution of N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
(100 mg, 0.16 mmol) in dichloromethane (5 mL) at 0 °C was added a solution of boron tribromide (0.5 mL, 5 mmol) in dichloromethane (5 mL). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated to give the crude product. The crude compound was purified by prep-TLC, using 5% methanol in dichloromethane as an eluent, to afford N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (Compound 26) as a brown solid (28 mg, 29%).
1H NMR (300 MHz, CD3OD): δ 8.46 (d, = 6.3 Hz, 2H), 7.87 (d, = 8.7 Hz, 2H), 7.53 (d, = 6.3 Hz, 2H), 7.38 (d, = 8.7 Hz, 1H), 7.12 (d, = 2.1 Hz, 1H), 6.97 (d, = 8.7 Hz, 2H), 6.90 (s, 2H), 6.83 (dd, = 8.7 Hz, 2.1 Hz, 1H), 4.21 (t, = 6.9 Hz, 2H), 2.54 (br s, 8H), 2.40-2.33 (m, 5H), 2.31 (s, 3H), 2.19 (s, 3H), 1.94 (quintet, = 6.9 Hz, 2H). LCMS: m/z 606.53 [M+H]+.
Other analogues prepared by this method:
Compound 27, N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(3 - (4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (19%). 1H NMR (300 MHz, CD3OD): δ 7.87 (d, = 8.7 Hz, 2H), 7.40-7.32 (m, 5H), 7.28-7.23 (m, 1H), 7.12 (d, J = 2.1 Hz, 1H), 6.97 (d, = 8.7 Hz, 2H), 6.87-6.81 (m, 3H), 4.20 (t, = 6.9 Hz, 2H), 2.65-2.32 (m, 16H), 2.19 (s, 3H), 1.94 (quintet, = 7.2 Hz, 2H). LCMS: m/z 605.57 [M+H]+.
Compound 28, 4-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)oxy)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4-yl)benzamide (7%).
1H NMR (400 MHz, CD3OD): δ 7.86 (d, = 8.8 Hz, 2H), 7.39-7.35 (m, 3H), 7.17 (d, J = 2.0 Hz, 1H), 7.07 (t, J = 8.8 Hz, 2H), 6.97 (d, J = 8.8 Hz, 2H), 6.86-6.81 (m, 3H), 4.21 (t, / = 6.8 Hz, 2H), 2.71-2.32 (m, 16H), 2.19 (s, 3H), 2.00-1.91 (m, 2H). LCMS: m/z 623.46 [M+H]+.
Scheme 15. Preparation of Compounds 30-32.
Figure imgf000092_0001
Preparation of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzonitrile
To a solution of 2,3-dimethyl- l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-ol (450 mg, 1.43 mmol) in degassed DMF (10 mL), was added K2C03 (0.99 g, 7.2 mmol) and 4-fluorobenzonitrile (0.51 g, 4.2 mmol) under an atmosphere of argon. The mixture was stirred at 120 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to 0 °C, quenched with ammonium chloride solution and extracted with ethyl acetate. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 3% methanol in dichloromethane as an eluent, to afford 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzonitrile as a yellow liquid (0.55 g, 92%). LCMS: m/z 417.31 [M+H]+.
Preparation of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoic acid
To a stirred solution of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzonitrile (0.55 g, 1.3 mmol) in EtOH (20 mL) was added 8 N KOH solution (5 mL) drop wise at 0 °C. The reaction mixture was heated to 100 °C as tirred at that temperature for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, acidified with 6 N HCl, and extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 4-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzoic acid as a brown solid (0.44 g, 76%). LCMS: m/z 436.32 [M+H]+.
Preparation of N-(2,6-dimethoxy-[l,l ' -biphenyl] -4-yl)-4-((2,3 -dimethyl-1 -(2- methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a stirred solution of 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzoic acid (80 mg, 0.18 mmol) and Intermediate C (69 mg, 0.30 mmol) in pyridine (3 mL), was added EDC.HC1 (211 mg, 1.1 mmol). The mixture was stirred at room temperature for 10 minutes, then heated to 100 °C for 2 hours. After complete consumption of the starting material, the reaction mixture was concentrated, diluted with aqueous ammonium chloride and extracted with ethyl acetate. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as the eluent, to afford N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4- ((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide as a brown solid (80 mg, 67%). LCMS: m/z 647.44 [M+H]+.
Other analogues prepared by this method: N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl -(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (46%). LCMS: m/z 647.01 [Μ-Η]".
4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)-N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (45%). LCMS: m/z 665.48 [M+H]+.
Preparation of Compound 31, N-(2,6-dihydroxy-[l,l '-biphenyl]-4-yl)-4-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide
To a stirred solution of N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)oxy)benzamide (80 mg, 0.12 mmol) in dichloromethane (6 mL) at 0 °C was added boron tribromide (2.0 mL, 2.0 mmol, 1.0 M in DCM). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. After complete consumption of the starting material, the reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by prep-TLC, using 8% methanol in DCM as an eluent, to afford N-(2,6-dihydroxy- [l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)oxy)benzamide (Compound 31) as a white solid (28 mg, 38%).
1H NMR (400 MHz, DMSO-d6): δ 9.94 (br s, 1H), 9.13 (br s, 2H), 7.92 (d, = 8.8 Hz, 2H), 7.41 (d, = 8.8 Hz, 1H), 7.31-7.30 (m, 4H), 7.22-7.18 (m, 1H), 7.15 (d, = 2.4 Hz, 1H), 6.97 (d, = 8.8 Hz, 2H), 6.94 (s, 2H) 6.83 (dd, = 8.0 Hz, 1.6 Hz, 1H), 4.19 (dd, = 14.8 Hz, 4.8 Hz, 1H), 3.88 (dd, / = 15.2 Hz, 9.2 Hz, 1H), 2.66-2.15 (m, 20H), 0.79 (d, = 6.4 Hz, 3H). LCMS: m/z 619.43 [M+H]+.
Other analogues prepared by this method:
Compound 30, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)oxy)benzamide (24%). 1H NMR (300 MHz, DMSO-d6): δ 10.01 (br s, 1H), 9.48 (s, 2H), 8.49 (d, = 6.0 Hz, 2H), 7.92 (d, = 8.8 Hz, 2H), 7.40 (d, = 8.8 Hz, 1H), 7.36 (d, = 6.0 Hz, 2H), 7.14 (d, = 2.4 Hz, 1H), 6.996.96 (m, 4H), 6.82 (dd, = 8.4 Hz, 2.4 Hz, 1H), 4.19 (dd, = 14.8 Hz, 4.4 Hz, 1H), 3.86 (dd, = 14.8 Hz, 8.8 Hz, 1H), 2.46-2.15 (m, 20H), 0.78 (d, J = 5.6 Hz, 3H). LCMS: m/z 620.31 [M+H]+.
Compound 32, 4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l- yl)propyl)- lH-indol-5-yl)oxy)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4- yl)benzamide (66%).
1H NMR (400 MHz, DMSO-d6): δ 9.94 (br s, 1H), 9.21 (br s, 2H), 7.91 (d, / = 8.8 Hz, 2H), 7.40 (d, = 8.8 Hz, 1H), 7.33 (dd, = 8.8 Hz, 6.0 Hz, 2H), 7.15-7.10 (m, 3H), 6.97 (d, J = 8.8 Hz, 2H), 6.94 (s, 2H), 6.82 (dd, J = 8.8 Hz, 2.0 Hz, 1H), 4.19 (dd, = 14.8 Hz, 4.4 Hz, 1H), 3.87 (dd, = 15.2 Hz, 8.0 Hz, 1H), 2.19-2.15 (m, 20H), 0.78 (d, J = 5.6 Hz, 3H). LCMS: m/z 637.39 [M+H]+.
Scheme 16. Preparation of Compounds 33-36 and 38-40.
Figure imgf000095_0001
Preparation of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- nitrobenzamide
To a stirred solution of 3-nitrobenzoic acid (350 mg, 2.1 mmol) in DMF (10 mL) at room temperature, DIPEA (1.06 mL, 6.09 mmol) was added, followed by the addition of HATU (1.5 g, 3.9 mmol) and then Intermediate A (505 mg, 2.3 mmol). The reaction mixture was stirred at room temperature for 16 hours. After complete consumption of the starting material, the reaction mass was diluted with ice water. The resulting precipitate was collected by filtration to give N-(4-(lH-imidazol-l-yl)- 3,5-dimethoxyphenyl)-3-nitrobenzamide as a brown solid (0.60 g, 78%).
1H NMR (400 MHz, DMSO-d6): δ 10.72 (br s, 1H), 8.83 (br s, 1H), 8.48-8.42 (m, 2H), 7.88 (t, 7 = 8.0 Hz, 1H), 7.57 (s, 1H), 7.40 (s, 2H), 7.12 (s, 1H), 7.00 (s, 1H), 3.76 (s, 6H). LCMS: m/z 369.1 [M+H]+.
Other analogues prepared by this method:
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-nitrobenzamide (82%).
1H NMR (300 MHz, DMSO-d6): δ 10.70 (br s, 1H), 8.84 (br s, 1H), 8.56 (d, J = 5.7 Hz, 2H), 8.49-8.42 (m, 2H), 7.88 (t, = 8.1 Hz, 1H), 7.36 (s, 2H), 7.28 (d, = 6.0 Hz, 2H), 3.72 (s, 6H). LCMS: m/z 380.1 [M+H]+.
N-(2,6-dimethoxy- [1,1 '-biphenyl] -4-yl)-3 -nitrobenzamide (72%) .
1H NMR (400 MHz, DMSO-d6): δ 10.64 (br s, 1H), 8.84 (t, = 2.0 Hz, 1H), 8.48-8.43 (m, 2H), 7.88 (t, = 8.0 Hz, 1H), 7.37-7.32 (m, 4H), 7.28-7.22 (m, 3H), 3.68 (s, 6H). LCMS: m/z 379.1 [M+H]+.
N-(4'-fluoro-2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-3-nitrobenzamide (84%).
1H NMR (400 MHz, DMSO-d6): δ 10.64 (br s, 1H), 8.83 (br s, 1H), 8.49-8.42 (m, 2H), 7.87 (t, = 8.0 Hz, 1H), 7.33 (s, 2H), 7.26 (dd, = 8.4 Hz, 5.6 Hz, 2H), 7.17 (t, = 8.8 Hz, 2H), 3.69 (s, 6H). LCMS: m/z 397.1 [M+H]+. Preparation of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- aminobenzamide
To a stirred solution of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- nitrobenzamide (600 mg, 1.8 mmol) in ethanol (30 mL) and water (30 mL), Fe powder (182 mg, 3.26 mmol) and NH4C1 (435 mg, 8.14 mmol) were added at room temperature. The reaction mixture was heated at reflux for 3 hours. After complete consumption of the starting material, the mixture was filtered through a plug of Celite and the filtrate concentrated under reduced pressure. The resulting precipitate was collected by filtration and dried to afford N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-3-aminobenzamide as a brown solid (400 mg, 73%).
1H NMR (400 MHz, DMSO-d6): δ 10.28 (br s, 1H), 7.58-7.34 (m, 3H), 7.27- 7.04 (m, 5H), 6.78 (d, J = 7.2 Hz, 1H), 3.76 (s, 6H). LCMS: m/z 339.1 [M+H]+.
Other analogues prepared by this method:
3-amino-N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)benzamide (99%).
1H NMR (400 MHz, DMSO-d6): δ 8.54 (br s, 2H), 7.37 (br s, 2H), 7.29 (br s, 2H), 7.21-7.05 (m, 4H), 6.77 (d, = 6.4 Hz, 1H), 5.40 (br s, 2H), 3.69 (s, 6H). LCMS: m/z 350.1 [M+H]+.
3-amino-N-(2,6-dimethoxy-[ 1 , 1 '-biphenyl]-4-yl)benzamide (72%).
1H NMR (400 MHz, DMSO-d6): δ 10.11 (br s, 1H), 7.35-7.32 (m, 4H), 7.26- 7.14 (m, 4H), 7.11-7.04 (m, 2H), 6.76 (br d, = 8.0 Hz, 1H), 5.35 (br s, 2H), 3.65 (s, 6H). LCMS: m/z 349.1 [M+H]+.
3-amino-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (500 mg,
77%).
1H NMR (400 MHz, DMSO-d6): δ 10.09 (br s, 1H), 7.33 (s, 2H), 7.26-7.22 (m, 2H), 7.17-7.07 (m, 5H), 6.76 (br d, = 8.0 Hz, 1H), 5.31 (br s, 2H), 3.72 (s, 6H). LCMS: m/z 367.1 [M+H]+. Preparation of 5-bromo-l-(3-chloropropyl)-2,3-dimethyl-lH-indole
NaH (1.8 g, 75 mmol) was added portionwise to a stirred solution of 5-bromo- 2,3-dimethyl-lH-indole (5.0 g, 22 mmol) in DMF (50 mL) at 0 °C. The mixture was allowed to warm to room temperature for 30 minutes. To this, bromochloropropane (11.7 mL, 11.8 mmol) was added dropwise at 0 °C and the mixture was allowed to stir at room temperature for 3 hours. After complete consumption of the starting material, ice-cold water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by flash column chromatography using 5% ethyl acetate in petroleum ether as an eluent to afford 5-bromo- l-(3-chloropropyl)-2,3-dimethyl-lH- indole as a pink solid (2.6 g, 39%).
1H NMR (400 MHz, CDC13): δ 7.59 (d, = 2.0 Hz, 1H), 7.21 (dd, = 8.4 Hz, 1.6 Hz, 1H), 7.16 (d, = 8.4 Hz, 1H), 4.22 (t, = 6.8 Hz, 2H), 3.49 (t, = 6.0 Hz, 2H), 2.36 (s, 3H), 2.30-2.15 (m, 5H). LCMS: m/z 301.2, 302.1 [M+H]+.
Other analogues prepared by this method:
5-bromo-l -(3 -chloro-2-methylpropyl)-2,3 -dimethyl- 1H- indole (61%).
1H NMR (400 MHz, CDCI3): δ 7.59 (d, = 2.0 Hz, 1H), 7.17 (dd, = 8.4 Hz, 2.0 Hz, 1H), 7.03 (d, = 8.4 Hz, 1H), 4.21-4.17 (m, 1H), 3.91-3.83 (m, 1H), 3.62- 3.49 (m, 2H), 2.36 (s, 3H), 2.30-2.25 (m, 4H), 1.02 (d, = 6.4 Hz, 3H). LCMS: m/z 315.1, 316.0 [M+H]+.
Preparation of 5-bromo-2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- IH-indole
To a stirred solution of 5-bromo-l-(3-chloropropyl)-2,3-dimethyl-lH-indole (9.0 g, 30 mmol) in acetonitrile (20 mL), sodium iodide (11.2 g, 74.7 mmol), sodium carbonate (7.93 g, 74.8 mmol) and then N-methylpiperazine (7.4 g, 73.9 mmol) were added at room temperature. The reaction mixture was heated to 75 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography using 5% methanol in dichloromethane as an eluent to afford 5-bromo-2,3-dimethyl- l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indole (3.2 g, 30%). LCMS: m/z 364.0, 366.0 [M+H]+.
Other analogues prepared by this method:
5-bromo-2,3-dimethyl- l-(2-methyl-3-(4-methylpiperazin- l-yl)propyl)- 1H- indole (17%).
1H NMR (400 MHz, CDC13): δ 7.58 (d, = 1.6 Hz, 1H), 7.17 (dd, = 8.4 Hz, 1.6 Hz, 1H), 7.11 (d, = 8.4 Hz, 1H), 4.22-4.17 (m, 1H), 3.76-3.70 (m, 1H), 2.65- 2.40 (m, 6H), 2.38 (s, 3H), 2.33 (s, 3H), 2.29-2.22 (m, 2H), 2.21-2.14 (m, 4H), 1.29- 1.25 (m, 2H), 0.87 (d, = 6.2 Hz, 3H). LCMS: m/z 378.1, 380.1 [M+H]+.
Preparation of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lll-indol-5-yl)amino)benzamide
To a stirred solution of 5-bromo-2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH- indole (200 mg, 0.55 mmol) in 1,4-dioxane (20 mL), under an argon atmosphere, was added NaOiBu (159 mg, 1.65 mmol), Pd2(dba)3 (56 mg, 0.061 mmol), Dave Phos (108 mg, 0.27 mmol) and N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-3-aminobenzamide (185 mg, 0.55 mmol) at room temperature. The reaction mixture was heated to 100 °C for 12 hours in a sealed tube. After complete consumption of the starting material, the reaction mixture was diluted with ethyl acetate and filtered through Celite. The organic layer was washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by prep-TLC using 5% methanol in dichloromethane as an eluent to afford N-(4-(lH-imidazol-l-yl)-3,5- dimethoxyphenyl)-3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol- 5-yl)amino)benzamide as a brown solid (80 mg, 23%). 1H NMR (400 MHz, DMSO-d6): δ 10.27 (br s, IH), 8.00 (br s, IH), 7.57-7.48 (m, IH), 7.43-7.39 (m, 3H), 7.35-7.17 (m, 4H), 7.11-7.05 (m, 2H), 7.00-6.93 (m, IH), 6.90 (br d, J = 8.0 Hz, IH), 4.10 (t, / = 6.8 Hz, 2H), 3.71 (s, 6H), 2.39-2.13 (m, 19H), 1.82-1.75 (m, 2H). LCMS: m/z 622.3 [M+H]+.
Other analogues prepared by this method:
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (99%).
1H NMR (400 MHz, DMSO-d6): δ 10.22 (br s, IH), 8.53 (d, = 5.2 Hz, 2H), 7.99 (br s, IH), 7.41 (br s, IH), 7.36-7.32 (m, 3H), 7.28-7.23 (m, 4H), 7.18 (br s, IH), 7.08 (br d, = 7.6 Hz, IH), 6.90 (d, = 8.0 Hz, IH), 4.10 (t, = 6.8 Hz, 2H), 3.68 (s, 6H), 2.38-2.20 (m, 13H), 2.15 (s, 6H), 1.79 (quintet, = 6.8 Hz, 2H). LCMS: m/z 633.3 [M+H]+.
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(3-(4- methylpiperazin- 1 -yl)propyl) - 1 H-indol- 5 -yl) amino)benzamide (26 % ) .
1H NMR (300 MHz, DMSO-d6): δ 10.16 (br s, IH), 7.98 (br s, IH), 7.41-7.18 (m, 10H), 7.08-6.89 (m, 4H), 4.10 (t, = 6.6 Hz, 2H), 3.64 (s, 6H), 2.41-2.15 (m, 19H), 1.81-1.74 (m, 2H). ). LCMS: m/z 632.4 [M+H]+.
3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)- N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (25%).
1H NMR (400 MHz, DMSO-d6): δ 10.18 (br s, IH), 7.96 (br s, IH), 7.41 (br s, IH), 7.34-7.29 (m, 3H), 7.27-7.21 (m, 4H), 7.18 (d, = 1.6 Hz, IH), 7.15 (t, = 9.2 Hz, 2H), 7.07 (br d, = 8.8 Hz, IH), 6.91 (dd, = 8.8 Hz, 2.0 Hz, IH), 4.10 (t, = 6.8 Hz, 2H), 3.65 (s, 6H), 2.38-2.14 (m, 19H), 1.79 (quintet, = 7.2 Hz, 2H). LCMS: m/z 650.3 [M+H]+.
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- 1 -yl)propyl) - 1 H-indol- 5 -yl) amino)benzamide (44 % ) . 1H NMR (300 MHz, DMSO-d6): δ 10.21 (br s, 1H), 8.54 (d, = 5.4 Hz, 2H), 7.99 (br s, 1H), 7.41 (br s, 1H), 7.37 (s, 2H), 7.36-7.18 (m, 6H), 7.10-7.04 (m, 1H), 6.88 (br d, J = 7.5 Hz, 1H), 6.90 (d, = 8.0 Hz, 1H), 4.19^1.11 (m, 1H), 3.87-3.76 (m, 1H), 3.68 (s, 6H), 2.50-2.04 (m, 20H), 0.78 (d, = 6.0 Hz, 3H). LCMS: m/z 647.6 [M+H]+.
N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (72%). LCMS: m/z 646.4 [M+H]+.
3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (26%).
1H NMR (400 MHz, DMSO-d6): δ 10.17 (br s, 1H), 7.98 (br s, 1H), 7.42 (br s, 1H), 7.31-7.22 (m, 7H), 7.18-7.13 (m, 3H), 7.07 (br d, = 7.2 Hz, 1H), 6.90 (br d, = 8.8 Hz, 1H), 4.19-4.12 (m, 1H), 3.86-3.78 (m, 1H), 3.65 (s, 6H), 2.41-2.12 (m, 20H), 0.93 (d, = 6.8 Hz, 3H). LCMS: m/z 664.3 [M+H]+.
Preparation of Compound 33, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)- 3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino )benzamide
To a stirred solution of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-3- ((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (50 mg, 0.080 mmol) in dichloromethane (3 mL) at 0 °C was added boron tribromide (1.0 mL, 1 M in dichloromethane). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. The reaction mixture was diluted with water and basified to pH 9 with saturated NaHC03. The resulting precipitate was collected by filtration to obtain the crude product. The crude compound was purified by prep-HPLC to afford N-(3,5-dihydroxy-4-(lH- imidazol-l-yl)phenyl)-3-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)amino)benzamide (Compound 33) as an off white solid (10 mg, 21%).
1H NMR (400 MHz, CD3OD): δ 8.47 (br s, 3H), 7.85 (d, = 8.4 Hz, 1H), 7.81 (d, = 8.0 Hz, 1H), 7.72 (br s, 1H), 7.48-7.41 (m, 1H), 7.38 (br s, 1H), 7.35-7.29 (m, 2H), 7.27-7.23 (m, 2H), 7.18-7.15 (m, 2H), 7.11-7.06 (m, 2H), 6.98 (dd, J = 8.4 Hz, 2.0 Hz, 1H), 6.93 (s, 2H), 4.19 (t, J = 6.8 Hz, 2H), 2.95-2.53 (m, 11H), 2.42-2.37 (m, 5H), 2.20 (s, 3H), 1.95 (quintet, J = 6.8 Hz, 2H). LCMS: m/z 594.40 [M+H]+.
Other analogues prepared by this method:
Compound 34, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (8%).
1H NMR (400 MHz, CD3OD): δ 8.46 (d, = 6.4 Hz, 2H), 7.54 (d, = 5.6 Hz, 2H), 7.38 (br s, 1H), 7.31 (d, = 8.8 Hz, 1H), 7.27-7.21 (m, 2H), 7.17 (br d, J = 7.2 Hz, 1H), 7.08 (br d, J = 7.6 Hz, 1H), 6.99 (dd, = 8.8 Hz, 1.6 Hz, 1H), 6.87 (s, 2H), 4.20 (t, = 6.8 Hz, 2H), 2.96 (br s, 8H), 2.64 (s, 3H), 2.42 (t, = 6.8 Hz, 2H), 2.37 (s, 3H), 2.20 (s, 3H), 1.96 (quintet, = 6.8 Hz, 2H). LCMS: m/z 605.46 [M+H]+.
Compound 35, N-(2,6-dihydroxy-[l, l'-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(3- (4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)amino)benzamide (12%).
1H NMR (400 MHz, CD3OD): δ 7.38-7.21 (m, 9H), 7.17 (br d, = 7.6 Hz, 1H), 7.08 (br d, = 9.2 Hz, 1H), 6.99 (dd, = 8.4 Hz, 2.0 Hz, 1H), 6.83 (s, 2H), 4.18 (t, = 6.8 Hz, 2H), 2.82-2.33 (m, 16H), 2.20 (s, 3H), 1.97-1.91 (m, 2H). LCMS: m/z 604.3 [M+H]+.
Compound 36, 3-((2,3-dimethyl-l-(3-(4-methylpiperazin- l-yl)propyl)- 1H- indol-5-yl)amino)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4-yl)benzamide (6%).
1H NMR (400 MHz, CD3OD): δ 7.39-7.34 (m, 3H), 7.29 (d, = 8.8 Hz, 1H), 7.26-7.22 (m, 2H), 7.17 (br s, = 8.0 Hz, 1H), 7.08-7.03 (m, 3H), 6.97 (dd, = 8.8 Hz, 2.0 Hz, 1H), 6.83 (s, 2H), 4.17 (t, = 6.8 Hz, 2H), 2.62-2.31 (m, 13H), 2.27 (s, 3H), 2.19 (s, 3H), 1.92 (quintet, = 6.8 Hz, 2H). LCMS: m/z 622.51 [M+H]+.
Compound 38, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-3-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (24%).
1H NMR (300 MHz, CD3OD): δ 8.46 (d, = 6.3 Hz, 2H), 7.53 (d, = 6.3 Hz, 2H), 7.39 (br s, 1H), 7.29-7.23 (m, 3H), 7.17 (br d, = 7.5 Hz, 1H), 7.09 (br d, = 6.6 Hz, IH), 6.98 (dd, J = 8.4 Hz, 2.1 Hz, IH), 6.87 (s, 2H), 4.19-4.13 (m, IH), 3.99- 3.92 (m, IH), 2.78 (br s, 8H), 2.59 (s, 3H), 2.39-2.32 (m, 6H), 2.21 (s, 3H), 0.93 (d, J = 5.7 Hz, 3H). LCMS: m/z 619.45 [M+H]+.
Compound 39, N-(2,6-dihydroxy-[l,r-biphenyl]-4-yl)-3-((2,3-dimethyl-l-(2- methyl-3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)amino)benzamide (17%).
1H NMR (300 MHz, DMSO-d6): δ 9.95 (br s, IH), 9.12 (br s, 2H), 8.17 (br s, IH), 7.96 (br s, IH), 7.39 (br s, IH), 7.31-7.16 (m, 8H), 7.05 (br d, J = 8.4 Hz, IH), 6.92 (s, 2H), 6.89 (dd, = 8.7 Hz, 1.8 Hz, IH), 4.19-4.11 (m, IH), 3.80 (dd, = 14.7 Hz, 8.4 Hz, IH), 2.45-2.13 (m, 20H), 0.77 (d, / = 6.0 Hz, 3H). LCMS: m/z 618.48 [M+H]+.
Compound 40, 3-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l- yl)propyl)-lH-indol-5-yl)amino)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4- yl)benzamide (3%).
1H NMR (400 MHz, CD3OD): δ 7.39-7.34 (m, 3H), 7.26-7.21 (m, 3H), 7.16 (br d, J = 6.4 Hz, IH), 7.09-7.03 (m, 3H), 6.96 (br d, = 8.8 Hz, IH), 6.83 (s, 2H), 4.20-4.16 (m, IH), 3.86 (dd, = 14.4 Hz, 5.6 Hz, IH), 2.53-2.32 (m, 11H), 2.35-2.25 (m, 6H), 2.15 (s, 3H), 0.88 (d, = 6.4 Hz, 3H). LCMS: m/z 636.3 [M+H]+.
Scheme 17. Preparation of Compounds 41-48.
Figure imgf000103_0001
Figure imgf000103_0002
Preparation ofN-(2,6-dimethoxy-[l,l '-biphenyl]-4-yl)-4-nitrobenzamide
To a stirred solution of 4-nitrobenzoic acid (350 mg, 2.1 mmol) in DMF (10 mL), HATU (1.5 g, 3.9 mmol), DIPEA (1.12 mL, 6.43 mmol) and Intermediate C (528 mg, 2.30 mmol) were added at room temperature. The reaction mixture was stirred at this temperature for 16 hours. After complete consumption of the starting material, the reaction mass was diluted with ice water. The resulting precipitate was collected by filtration to give N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4- nitrobenzamide as an off white solid (600 mg, 76%).
1H NMR (400 MHz, DMSO-d6): δ 10.62 (br s, 1H), 8.40 (d, / = 8.8 Hz, 2H), 8.22 (d, J = 8.8 Hz, 2H), 7.38-7.31 (m, 4H), 7.28-7.21 (m, 3H), 3.67 (s, 6H). LCMS: m/z 379.0 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-nitrobenzamide (73%).
1H NMR (400 MHz, DMSO-d6): δ 10.72 (br s, 1H), 8.41 (d, = 8.8 Hz, 2H), 8.22 (d, J = 8.8 Hz, 2H), 7.57 (s, 1H), 7.39 (s, 2H), 7.12 (s, 1H), 6.99 (s, 1H), 3.75 (s, 6H). LCMS: m/z 369.1 [M+H]+.
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-nitrobenzamide (56%). LCMS: m/z 380.1 [M+H]+.
N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-nitrobenzamide (84%).
1H NMR (300 MHz, DMSO-d6): δ 10.61 (br s, 1H), 8.40 (d, = 8.7 Hz, 2H), 8.21 (d, = 8.7 Hz, 2H), 7.32 (s, 2H), 7.26 (dd, = 8.7 Hz, 6.0 Hz, 2H), 7.17 (t, = 9.0 Hz, 2H), 3.68 (s, 6H). LCMS: m/z 397.1 [M+H]+.
Preparation of4-amino-N-(2,6-dimethoxy-[l,l '-biphenyl] -4-yl)benzamide
To a stirred solution of N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4- nitrobenzamide (600 mg, 1.58 mmol) in ethanol (15 mL) and water (15 mL), Fe powder (177 mg, 3.17 mmol) and NH4C1 (423 mg, 7.91 mmol) were added at room temperature. The reaction mixture was heated at reflux for 3 hours. After complete consumption of the starting material, the mixture was filtered through a plug of Celite and the filtrate concentrated under reduced pressure. The resulting precipitate was collected by filtration and dried to afford 4-amino-N-(2,6-dimethoxy-[ 1,1 '-biphenyl] - 4-yl)benzamide as an off white solid (400 mg, 72%). LCMS: m/z 349.1 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-aminobenzamide (73%). LCMS: m/z 339.1 [M+H]+.
4-amino-N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)benzamide (81%). LCMS: m/z 350.1 [M+H]+.
4-amino-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (400 mg,
62%).
1H NMR (400 MHz, DMSO-d6): δ 9.80 (br s, 1H), 7.74 (d, = 7.6 Hz, 2H), 7.32 (s, 2H), 7.25-7.13 (m, 4H), 6.61 (d, = 8.0 Hz, 2H), 5.79 (br s, 2H), 3.66 (s, 6H). LCMS: m/z 367.1 [M+H]+.
Preparation of 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH- indol-5-yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l,l '-biphenyl] -4-yl)benzamide
To a solution of 5-bromo-2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indole (200 mg, 0.55 mmol) in 1,4-dioxane (20 mL) was added NaOiBu (159 mg, 1.65 mmol), Pd2(dba)3 (56 mg, 0.061 mmol), Dave Phos (108 mg, 0.27 mmol) and 4-amino-N-(4'-fluoro-2,6-dimethoxy- [1,1 '-biphenyl] -4-yl)benzamide (200 mg, 0.55 mmol) at room temperature. The reaction mixture was heated to 100 °C for 12 hours in a sealed tube. After complete consumption of the starting material, the reaction mixture was diluted with ethyl acetate and filtered through a plug of Celite. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by prep-TLC, using 5% methanol in dichloromethane as an eluent to afford 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide as a brown gummy solid (120 mg, 34%).
1H NMR (400 MHz, DMSO-d6): δ 9.90 (br s, 1H), 8.38 (br s, 1H), 7.83 (d, J = 8.8 Hz, 2H), 7.38-7.33 (m, 3H), 7.26-7.20 (m, 3H), 7.16 (t, J = 8.8 Hz, 2H), 6.94- 6.91 (m, 3H), 4.11 (t, J = 6.4 Hz, 2H), 3.66 (s, 6H), 2.46-2.15 (m, 19H), 1.83-1.75 (m, 2H). LCMS: m/z 650.2 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol- l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl-l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide (21%).
1H NMR (300 MHz, DMSO-d6): δ 9.98 (br s, 1H), 8.39 (br s, 1H), 7.85 (d, J = 8.7 Hz, 2H), 7.54 (s, 1H), 7.42 (s, 2H), 7.36 (d, = 8.7 Hz, 1H), 7.21 (br s, 1H), 7.09 (s, 1H), 6.98 (s, 1H), 6.95-6.90 (m, 3H), 4.11 (t, J = 6.6 Hz, 2H), 3.72 (s, 6H), 2.43- 2.14 (m, 19H), 1.85-1.73 (m, 2H). LCMS: m/z 622.2 [M+H]+.
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide (23%).
1H NMR (300 MHz, DMSO-d6): δ 9.94 (br s, 1H), 8.53 (d, = 5.4 Hz, 2H), 8.38 (br s, 1H), 7.83 (d, = 8.7 Hz, 2H), 7.40-7.35 (m, 3H), 7.27 (d, = 5.7 Hz, 2H), 7.21 (br s, 1H), 6.96-6.89 (m, 3H), 4.11 (t, J = 6.9 Hz, 2H), 3.69 (s, 6H), 2.48-2.14 (m, 19H), 1.84-1.72 (m, 2H). LCMS: m/z 633.5 [M+H]+.
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl- l-(3-(4- methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide (23%).
1H NMR (300 MHz, DMSO-d6): δ 9.88 (br s, 1H), 8.36 (br s, 1H), 7.83 (d, J = 8.7 Hz, 2H), 7.38-7.31 (m, 5H), 7.27-7.19 (m, 4H), 6.95-6.89 (m, 3H), 4.11 (t, J = 6.6 Hz, 2H), 3.65 (s, 6H), 2.45-2.14 (m, 19H), 1.80 (quintet, J = 6.6 Hz, 2H). LCMS: m/z 632.4 [M+H]+.
N-(4-(lH-imidazol- l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl-l-(2-methyl- 3-(4-methylpiperazin- l-yl)propyl)-lH-indol-5-yl)amino)benzamide (20%). 1H NMR (300 MHz, DMSO-d6): δ 9.99 (br s, IH), 8.38 (br s, IH), 7.83 (d, 7 = 8.7 Hz, 2H), 7.54 (br s, IH), 7.42 (s, 2H), 7.32 (d, 7 = 8.4 Hz, IH), 7.21 (br s, IH), 7.09 (br s, IH), 6.98-6.89 (m, 4H), 4.22-4.14 (m, IH), 3.88-3.76 (m, IH), 3.72 (s, 6H), 2.42-2.14 (m, 20H), 0.78 (d, 7 = 6.0 Hz, 3H). LCMS: m/z 636.4 [M+H]+.
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- 1 -yl)propyl) - 1 H-indol- 5 -yl) amino)benzamide (16%).
1H NMR (400 MHz, DMSO-d6): δ 9.94 (br s, IH), 8.55 (d, 7 = 5.6 Hz, 2H), 8.38 (br s, IH), 7.82 (d, 7 = 8.8 Hz, 2H), 7.39 (s, 2H), 7.36 (d, 7 = 8.4 Hz, IH), 7.28 (d, 7 = 5.6 Hz, 2H), 7.22 (d, 7 = 2.0 Hz, IH), 6.97-6.92 (m, 3H), 4.20-4.14 (m, IH), 3.86-3.79 (m, IH), 3.69 (s, 6H), 2.49-2.05 (m, 20H), 0.79 (d, 7 = 6.4 Hz, 3H). LCMS: m/z 647.4 [M+H]+.
N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-methyl-3-(4- methylpiperazin- 1 -yl)propyl) - 1 H-indol- 5 -yl) amino)benzamide (35%).
1H NMR (300 MHz, DMSO-d6): δ 9.88 (br s, IH), 8.35 (br s, IH), 7.83 (d, 7 =
8.7 Hz, 2H), 7.40-7.28 (m, 5H), 7.27-7.19 (m, 4H), 6.96-6.89 (m, 3H), 4.17 (dd, 7 = 14.7 Hz, 4.2 Hz, IH), 3.83 (dd, 7 = 13.5 Hz, 8.1 Hz, IH), 3.65 (s, 6H), 2.46-2.12 (m, 20H), 0.78 (d, 7 = 6.0 Hz, 3H). LCMS: m/z 646.4 [M+H]+.
4-((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (34%).
1H NMR (400 MHz, DMSO-d6): δ 9.89 (br s, IH), 8.36 (br s, IH), 7.83 (d, 7 =
8.8 Hz, 2H), 7.39-7.30 (m, 3H), 7.26-7.21 (m, 3H), 7.16 (t, 7 = 8.8 Hz, 2H), 6.94- 6.90 (m, 3H), 4.16 (dd, 7 = 14.8 Hz, 4.4 Hz, IH), 3.83 (dd, 7 = 14.8 Hz, 8.4 Hz, IH), 3.66 (s, 6H), 2.46-2.15 (m, 20H), 0.78 (d, 7 = 6.4 Hz, 3H). LCMS: m/z 664.3 [M+H]+.
Preparation of Compound 44, 4-((2,3-dimethyl-l-(3-(4-methylpiperazin-l- yl)propyl)-lH-indol-5-yl)amino)-N-(4'-fluoro-2,6-dihydroxy-[l,l '-biphenyl]-4- yl)benzamide To a stirred solution of 4-((2,3-dimethyl- l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)amino)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (120 mg, 0.18 mmol) in dichloromethane (3 mL) at 0 °C was added boron tribromide (1.0 mL, 1 M in dichloromethane). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. The reaction mixture was then concentrated, diluted with water and then basified to pH 9 with saturated NaHC03. The resulting precipitate was collected by filtration to give the crude product. The crude compound was purified by prep-HPLC to afford 4-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)amino)-N-(4'-fluoro- 2,6-dihydroxy-[l,l'-biphenyl]-4-yl)benzamide (Compound 44) as a brown solid (20 mg, 18%).
1H NMR (400 MHz, CD3OD ): δ 7.74 (d, = 8.8 Hz, 2H), 7.37 (dd, = 8.8 Hz, 5.6 Hz, 2H), 7.31 (d, = 8.8 Hz, 1H), 7.25 (d, = 2.0 Hz, 1H), 7.06 (t, = 8.8 Hz, 2H), 6.97 (dd, = 8.4 Hz, 2.0 Hz, 1H), 6.91 (d, = 8.8 Hz, 2H), 6.83 (s, 2H), 4.18 (t, = 6.8 Hz, 2H), 2.68-2.30 (m, 16H), 2.20 (s, 3H), 1.94 (quintet, = 7.2 Hz, 2H). LCMS: m/z 622.3 [M+H]+.
Other analogues prepared by this method:
Compound 41, N-(3,5-dihydroxy-4-(lH-imidazol- l-yl)phenyl)-4-((2,3- dimethyl-l-(3-(4-methylpiperazin-l-yl)propyl)- lH-indol-5-yl)amino)benzamide (4%).
1H NMR (400 MHz, DMSO-d6): δ 9.78 (br s, 3H), 8.33 (br s, 1H), 7.79 (d, = 8.0 Hz, 2H), 7.53 (br s, 1H), 7.35 (s, = 8.8 Hz, 1H), 7.20 (br s, 1H), 7.09 (s, 1H), 7.04 (s, 2H), 6.95-6.89 (m, 4H), 4.12 (br s, 2H), 2.49-2.14 (m, 19H), 1.87-1.72 (m, 2H). ). LCMS: m/z 594.3 [M+H]+.
Compound 42, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (6%).
1H NMR (400 MHz, CD3OD): δ 8.46 (d, = 5.6 Hz, 2H), 7.74 (d, = 8.8 Hz, 2H), 7.54 (d, = 6.0 Hz, 2H), 7.31 (d, = 8.8 Hz, 1H), 7.25 (br s, 1H), 6.97 (br d, = 8.8 Hz, 1H), 6.92 (d, = 8.8 Hz, 2H), 6.88 (s, 2H), 4.18 (t, = 7.2 Hz, 2H), 2.65-2.33 (m, 13H), 2.30 (s, 3H), 2.20 (s, 3H), 1.94 (quintet, J = 7.2 Hz, 2H). LCMS: m/z 605.3 [M+H]+.
Compound 43, N-(2,6-dihydroxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(3- (4-methylpiperazin- l-yl)propyl)- lH-indol-5-yl)amino)benzamide (26%).
1H NMR (400 MHz, DMSO-d6): δ 9.66 (br s, IH), 9.08 (br s, 2H), 8.30 (br s, IH), 7.79 (d, = 8.4 Hz, 2H), 7.37-7.08 (m, 7H), 6.94-6.90 (m, 5H), 4.11 (t, = 6.8 Hz, 2H), 2.50-2.14 (m, 19H), 1.86-1.74 (m, 2H). LCMS: m/z 604.4 [M+H]+.
Compound 45, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)-4-((2,3- dimethyl-l-(2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5- yl)amino)benzamide (4%).
1H NMR (400 MHz, CD3OD): δ 7.74 (d, = 8.8 Hz, 2H), 7.68 (br s, IH), 7.29-7.25 (m, 2H), 7.14 (br s, IH), 7.06 (br s, IH), 6.98-6.90 (m, 5H), 4.20 (dd, = 16.0 Hz, 5.2 Hz, IH), 3.91 (dd, = 15.2 Hz, 8.0 Hz, IH), 2.69-2.44 (m, 11H), 2.40- 2.28 (m, 6H), 2.21 (s, 3H), 0.90 (d, = 5.6 Hz, 3H). LCMS: m/z 606.4 [M-H]~.
Compound 46, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (2-methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (6%).
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 6.0 Hz, 2H), 7.75 (d, = 8.8 Hz, 2H), 7.54 (d, = 5.6 Hz, 2H), 7.28-7.25 (m, 2H), 6.96 (dd, = 8.8 Hz, 2.0 Hz, IH), 6.92 (d, = 8.8 Hz, 2H), 6.88 (s, 2H), 4.15 (dd, = 13.6 Hz, 4.0 Hz, IH), 3.90 (dd, = 14.0 Hz, 8.0 Hz, IH), 2.65-2.39 (m, 8H), 2.37 (s, 3H), 2.35 (s, 3H), 2.32-2.26 (m, 3H), 2.21 (s, 3H), 0.89 (d, = 5.6 Hz, 3H). LCMS: m/z 617 '.4 [M-H]".
Compound 47, N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(2- methyl-3-(4-methylpiperazin-l-yl)propyl)-lH-indol-5-yl)amino)benzamide (22%).
1H NMR (400 MHz, CD3OD): δ 7.75 (d, = 8.8 Hz, 2H), 7.38-7.31 (m, 4H), 7.28-7.22 (m, 3H), 6.96 (dd, = 8.0 Hz, 2.0 Hz, IH), 6.92 (d, = 8.8 Hz, 2H), 6.84 (s, 2H), 4.20 (dd, = 14.4 Hz, 5.2 Hz, IH), 3.80 (dd, = 14.8 Hz, 6.8 Hz, IH), 2.62- 2.38 (m, 8H), 2.37 (s, 3H), 2.33 (s, 3H), 2.31-2.26 (m, 3H), 0.89 (d, J = 6.0 Hz, 3H). LCMS: m/z 616.4 [M-H]~.
Compound 48, 4((2,3-dimethyl-l-(2-methyl-3-(4-methylpiperazin- l- yl)propyl)-lH-indol-5-yl)amino)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4- yl)benzamide (22%).
1H NMR (400 MHz, CD3OD): δ 7.74 (d, = 8.8 Hz, 2H), 7.37 (dd, = 8.8 Hz, 6.0 Hz, 2H), 7.28-7.24 (m, 2H), 7.06 (t, = 8.8 Hz, 2H), 6.96 (dd, = 8.8 Hz, 2.4 Hz, 1H), 6.92 (d, = 8.8 Hz, 2H), 6.83 (s, 2H), 4.20 (dd, = 14.8 Hz, 4.8 Hz, 1H), 3.87 (dd, / = 14.8 Hz, 8.4 Hz, 1H), 2.56-2.34 (m, 11H), 2.33-2.25 (m, 6H), 2.21 (s, 3H), 0.88 (d, J = 6.0 Hz, 3H). LCMS: m/z 634.4 [M-H]\
Scheme 18. Preparation of Compounds 57-60.
Figure imgf000110_0001
Preparation of5-bromo-l-(2-chloroethyl)-2,3-dimethyl-lH-indole
To a solution of 5-bromo-2,3-dimethyl-lH-indole (3.0 g, 13 mmol) in dimethyl sulfoxide (30 mL) was added ΚΟΗ (3.1 g, 55 mmol) at 0 °C. The mixture was allowed to warm to room temperature and was stirred for 20 minutes before being cooled to 0 °C. 2-Chloroethyl 4-methylbenzenesulfonate (9.0 g, 38 mmol) was added dropwise. The mixture was allowed to warm to room temperature and stirred for 16 hours. After complete consumption of the starting material, ice-cold water was added and the reaction mixture was extracted with ethyl acetate. The organic layer was washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 1% ethyl acetate in petroleum ether as an eluent, to afford 5-bromo-l-(2-chloroethyl)-2,3- dimethyl-lH-indole as a yellow solid (1.28 g, 33%). LCMS: m/z 285.9, 287.9 [M+H]+.
Preparation of 5-bromo-2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH- indole
To a stirred solution of 5-bromo-l-(2-chloroethyl)-2,3-dimethyl-lH-indole (5.2 g, 18 mmol) in acetonitrile (50 mL), sodium iodide (8.2 g, 55 mmol), sodium carbonate (5.7 g, 54 mmol) and then N-methylpiperazine (5.4 g, 54 mmol) were added at room temperature. The reaction mixture was stirred at 90 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was quenched with ammonium chloride solution and extracted with ethyl acetate. The organic layers were washed with aqueous ammonium chloride and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 6% methanol in dichloromethane as the eluent, to afford 5-bromo-2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indole as a yellow semi-solid (3.3 g, 52%). LCMS: m/z 350.36, 352.36 [M+H]+.
Preparation of 2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indole-5- carbaldehyde
To a stirred solution of 5-bromo-2,3-dimethyl-l-(2-(4-methylpiperazin-l- yl)ethyl)-lH-indole (3.3 g, 9.4 mmol), in dry THF (45 mL) was added n-BuLi (11.3 mL, 28.5 mmol, 2.5 M in THF) dropwise at -78 °C. Then dry DMF (3.0 mL) was added to the reaction mass and the resulting reaction mixture was stirred at -78 °C for 30 minutes. After complete consumption of the starting material, the reaction mass was quenched with saturated ammonium chloride solution and extracted with ethyl acetate. The combined organic layers were washed with aqueous ammonium chloride and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford crude 2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)- lH-indole-5-carbaldehyde as a yellow semi solid (3.1 g, crude), which was used in the subsequent reaction without any further purification. LCMS: m/z 300.3 [M+H]+.
Preparation of N'-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH- indol-5-yl)methylene)-4-methylbenzenesulfonohydrazide
Tosyl hydrazide (3.3 g, 18 mmol) was added to a stirred solution of
2.3- dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indole-5-carbaldehyde (3.5 g, 12 mmol) in dry 1,4-dioxane (80 mL) at room temperature. The mixture was stirred at room temperature for 4 hours. The crude N'-((2,3-dimethyl-l-(2-(4-methylpiperazin- l-yl)ethyl)-lH-indol-5-yl)methylene)-4-methylbenzenesulfonohydrazide in the reaction mixture was used without isolation or further purification..
Preparation of methyl 4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)- lH-indol-5-yl)methyl)benzoate
To the solution of crude N'-((2,3-dimethyl-l-(2-(4-methylpiperazin-l- yl)ethyl)- lH-indol-5-yl)methylene)-4-methylbenzenesulfonohydrazide in dry
1.4- dioxane (80 mL) was added K2C03 (4.8 g, 25 mmol) and (4-(methoxycarbonyl)phenyl)boronic acid (3.16 g, 17.6 mmol). The reaction temperature was raised to 110 °C and maintained for 5 hours. After complete consumption of the starting material, the reaction mass was quenched with water and extracted with ethyl acetate. The combined organic layers were washed with aqueous ammonium chloride and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as an eluent, to obtain methyl 4-((2,3-dimethyl-l-(2-(4- methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzoate as a yellow liquid (2.2 g, 44%). LCMS: m/z 420.38 [M+H]+. Preparation of 4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indol- 5-yl)methyl)benzoic acid
To a stirred solution of methyl 4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l- yl)ethyl)-lH-indol-5-yl)methyl)benzoate (2.1 g, 5.0 mmol) in THF:MeOH (4: 1, 30 mL) was added aqueous 2 N NaOH solution (20 mL) dropwise at 0 °C. The reaction mixture was allowed to warm to room temperature and was stirred for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated and then partitioned between ethyl acetate and water. The aqueous layer was acidified to pH 2 with 2 N HC1 and extracted with 30% isopropyl alcohol in chloroform. The combined organic layers were was with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 4-((2,3-dimethyl-l-(2-(4- methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzoic acid as an off white solid (1.1 g, 52%). LCMS: m/z 406.38 [M+H]+.
Preparation of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3- dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-l i-indol-5-yl)methyl)benzamide
To a solution of 4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH- indol-5-yl)methyl)benzoic acid (250 mg, 0.62 mmol) and Intermediate A (175 mg, 0.80 mmol) in pyridine (4 mL), was added EDC.HC1 (710 mg, 3.7 mmol). The mixture was stirred at room temperature for ten minutes, then at 80 °C for one hour. The reaction mixture was quenched with aqueous ammonium chloride and extracted with ethyl acetate. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichloromethane as an eluent, to afford N-(4- (lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3 -dimethyl- 1 -(2-(4-methylpiperazin- l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide as a brown solid (230 mg, 62%). LCMS: m/z 607.56 [M+H]+.
Other analogues prepared by this method: N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1 -(2-(4- methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide (47%). LCMS: m/z 616.49 [Μ-Η]".
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(2-(4- methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide (52%). LCMS: m/z 617.47 [M+H]+.
4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)- N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (51%). LCMS: m/z 635.47 [M+H]+.
Preparation of Compound 57, N-(3,5-dihydroxy-4-(lH-imidazol-l-yl)phenyl)- 4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lli-indol-5-yl)methyl)benzamide
To a stirred solution of N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4- ((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide (230 mg, 0.38 mmol) in dichloromethane (5 mL) at 0 °C was added a solution of boron tribromide (0.50 mL, 5.3 mmol) in dichloromethane (5 mL). The mixture was stirred at this temperature for one hour, then allowed to warm to room temperature and stirred for 16 hours. After complete consumption of the starting material, the reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layer was washed with brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to obtain the crude product. The crude compound was purified by prep-TLC, using 7% methanol in dichloromethane as an eluent, to afford N-(3,5-dihydroxy-4-(lH-imidazol-l- yl)phenyl)-4-((2,3-dimethyl-l-(2-(4-methylpiperazin-l-yl)ethyl)-lH-indol-5- yl)methyl)benzamide (Compound 57) as a brown solid (55 mg, 25%).
1H NMR (400 MHz, CD3OD): δ 7.80 (d, 7 = 8.0 Hz, 2H), 7.65 (br s, 1H), 7.36 (d, = 8.4 Hz, 2H), 7.25 (br s, 1H), 7.22 (d, = 8.8 Hz, 1H), 7.13 (br s, 1H), 7.05 (br s, 1H), 6.97-6.93 (m, 3H), 4.22 (t, = 6.8 Hz, 2H), 4.12 (s, 2H), 2.64-2.43 (m, 10H), 2.36 (s, 3H), 2.32 (s, 3H), 2.18 (s, 3H). LCMS: m/z 579.45 [M+H]+.
Other analogues prepared by this method: Compound 58, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (2-(4-methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide (42%).
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 6.4 Hz, 2H), 7.80 (d, = 8.4 Hz, 2H), 7.52 (d, = 6.0 Hz, 2H), 7.35 (d, = 8.4 Hz, 2H), 7.25 (br s, 1H), 7.21 (d, = 8.4 Hz, 1H), 6.95 (dd, = 8.4 Hz, 1.6 Hz, 1H), 6.90 (s, 2H), 4.21 (t, = 7.2 Hz, 2H), 4.11 (s, 2H), 2.65-2.49 (m, 10H), 2.36 (s, 3H), 2.32 (s, 3H), 2.18 (s, 3H). LCMS: m/z 588.56 [M-H]~.
Compound 59, N-(2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(2- (4-methylpiperazin-l-yl)ethyl)-lH-indol-5-yl)methyl)benzamide (16%).
1H NMR (400 MHz, CD3OD): δ 7.80 (d, = 8.0 Hz, 2H), 7.38-7.32 (m, 6H), 7.26-7.20 (m, 3H), 6.95 (dd, / = 8.4 Hz, 1.6 Hz, 1H), 6.86 (s, 2H), 4.22 (t, / = 7.2 Hz, 2H), 4.11 (s, 2H), 2.65-2.42 (m, 10H), 2.36 (s, 3H), 2.31 (s, 3H), 2.18 (s, 3H).
LCMS: m/z 589.48 [M+H]+.
Compound 60, 4-((2,3-dimethyl- l-(2-(4-methylpiperazin-l-yl)ethyl)- 1H- indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy-[ 1,1 '-biphenyl] -4-yl)benzamide (29%).
1H NMR (400 MHz, CD3OD): δ 7.80 (d, = 7.6 Hz, 2H), 7.39-7.34 (m, 4H), 7.25 (br s, 1H), 7.21 (d, = 8.0 Hz, 1H), 7.06 (t, = 8.8 Hz, 2H), 6.94 (dd, = 8.4 Hz, 1.6 Hz, 1H), 6.85 (s, 2H), 4.21 (t, = 7.2 Hz, 2H), 4.11 (s, 2H), 2.62-2.40 (m, 10H), 2.36 (s, 3H), 2.28 (s, 3H), 2.18 (s, 3H). LCMS: m/z 607.16 [M+H]+.
Scheme 19. Preparation of Compounds 61-68
Figure imgf000116_0001
61 : n = 4 i X = 1 -imidazolyl 65: n = 5, X = 1 -imidazolyl
62: n = 4 χ = -pyrjdyi 66: n = 5, = 4-pyridyl
63: n = 4i x = pheny| 67: n = 5, X = phenyl
64: n = 4 x = 4-fluorophenyl 68: n = 5, X = 4-fluorophenyl
Preparation of 4-((l-(4-chlorobutyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoic acid
A suspension of NaH (0.32 g, 13 mmol) in dry DMF (5 mL) was added portionwise to a stirred solution of methyl 4-((2,3-dimethyl-lH-indol-5- yl)methyl)benzoate (0.80 g, 2.7 mmol) in DMF (5 mL) at 0 °C. The mixture was stirred at this temperature for 30 minutes. l-Bromo-4-chlorobutane (1.0 mL, 8.7 mmol) was added dropwise at 0 °C and the mixture was stirred for one hour. After complete consumption of the starting material, the reaction was quenched with saturated ammonium chloride solution, diluted with ice-cold water and extracted with ethyl acetate. The organic layer was washed with brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford crude 4-((l-(4- chlorobutyl)-2,3-dimethyl-lH-indol-5-yl)methyl)benzoic acid as a brown liquid (0.8 g, crude), which was used in the subsequent reaction without further purification. LCMS: m/z 370.1 [M+H]+.
Other analogues prepared by this method: 4-((l-(5-chloropentyl)-2,3-dimethyl- lH-indol-5-yl)methyl)benzoic acid (80%, crude). LCMS: m/z 384.1 [M+H]+.
Preparation of methyl 4-((l-(4-chlorobutyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoate
To a solution of 4-((l-(4-chlorobutyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoic acid (0.80 g, 2.2 mmol) in methanol (20 mL) was added concentrated sulphuric acid (0.5 mL) dropwise at 0 °C. The mixture was heated to 80 °C for 16 hours. Afte complete consumption of the starting material, the reaction mixture was concentrated, then diluted with ethyl acetate. The organic layer was washed with saturated NaHC03 solution, water and brine, dried over anhydrous Na2S04 and evaporated in vacuo to give methyl 4-((l-(4-chlorobutyl)-2,3-dimethyl- lH-indol-5-yl)methyl)benzoate as a brown liquid (0.8 g, crude), which was used in the subsequent reaction without purification. LCMS: m/z 384.1 [M+H]+.
Other analogues prepared by this method:
4-((l-(5-chloropentyl)-2,3-dimethyl-lH-indol-5-yl)methyl)benzoate (81%, crude). LCMS: m/z 398.1 [M+H]+.
Preparation of methyl 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)- lH-indol-5-yl)methyl)benzoate
To a stirred solution of methyl 4-((l-(4-chlorobutyl)-2,3-dimethyl-lH-indol-5- yl)methyl)benzoate (0.80 g, 2.1 mmol) in acetonitrile (20 mL), sodium iodide (0.78 g, 5.2 mmol), sodium carbonate (0.40 g, 3.8 mmol) and then N-methylpiperazine (0.38 mL, 3.4 mmol) were added at room temperature. The reaction mixture was heated to 85 °C for 16 hours. After complete consumption of the starting material, the reaction mixture was cooled to room temperature and concentrated. The residue was diluted with ethyl acetate, washed with water and brine solution, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford the crude product. The crude compound was purified by flash column chromatography using 100-200 mesh silica gel, with 4% methanol in dichloromethane as an eluent, to afford methyl 4- ((2,3-dimethyl- l-(4-(4-methylpiperazin- l-yl)butyl)- lH-indol-5-yl)methyl)benzoate as a yellow liquid (0.56 g, 60%).
1H NMR (400 MHz, CDC13): δ 7.93 (d, = 8.0 Hz, 2H), 7.28 (d, = 8.0 Hz, 2H), 7.25 (br s, 1H), 7.15 (d, = 8.0 Hz, 1H), 6.92 (br d, J = 9.6 Hz, 1H), 4.12 (s, 2H), 4.03 (t, = 7.2 Hz, 2H), 3.89 (s, 3H), 2.63-2.22 (m, 16H), 2.20 (s, 3H), 1.75- 1.57 (m, 4H). LCMS: m/z 448.3 [M+H]+.
Other analogues prepared by this method: methyl 4-((2,3-dimethyl-l-(5-(4-methylpiperazin-l-yl)pentyl)-lH-indol-5- yl)methyl)benzoate (66%).
1H NMR (400 MHz, CDC13): δ 7.93 (d, = 8.0 Hz, 2H), 7.28 (d, = 8.0 Hz, 2H), 7.25 (br s, 1H), 7.13 (d, = 8.4 Hz, 1H), 6.92 (br d, = 9.2 Hz, 1H), 4.12 (s, 2H), 4.02 (t, = 7.2 Hz, 2H), 3.89 (s, 3H), 2.99-2.41 (m, 11H), 2.32 (s, 3H), 2.21 (s, 3H), 1.75-1.70 (m, 2H), 1.55-1.46 (m, 4H), 1.37-1.25 (m, 2H). LCMS: m/z 462.3 [M+H]+.
Preparation of '4-((2,3 -dimethyl-1 -(4-(4-methylpiperazin-l -yl)butyl)-lH-indol- 5-yl)methyl)benzoic acid
To a solution of methyl 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)- lH-indol-5-yl)methyl)benzoate (0.56 g, 1.3 mmol) in THF:MeOH (4: 1, 10 mL) was added 2 N NaOH solution (5 mL) at 0 °C. The reaction mixture was allowed to warm to room temperature and was stirred for 16 hours. After complete consumption of the starting material, the reaction mass was concentrated, neutralised with 2 N HC1 and extracted with 30% isopropyl alcohol in dichloromethane. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to afford 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)-lH- indol-5-yl)methyl)benzoic acid as a brown sold (0.46 g, 85%). LCMS: m/z 434.2 [M+H]+.
Other analogues prepared by this method: 4-((2,3-dimethyl-l-(5-(4-methylpiperazin-l-yl)pentyl)-lH-indol-5- yl)methyl)benzoic acid (93%). LCMS: m/z 448.3 [M+H]+.
Preparation of 4-((2,3 -dimethyl-1 -(4-(4-methylpiperazin-l -yl)butyl)-lH-indol- 5-yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide
To a solution of 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)-lH- indol-5-yl)methyl)benzoic acid (150 mg, 0.35 mmol) and Intermediate D (113 mg, 0.46 mmol) in pyridine (3 mL), was added EDC.HCl (325 mg, 2.1 mmol). The mixture was stirred at room temperature for 10 minutes, then at 80 °C for one hour. After complete consumption of the starting material, The reaction mass was concentrated, diluted with aqueous ammonium chloride and extracted with ethyl acetate. The organic layer was washed with water and brine, dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by column chromatography on 100-200 mesh silica gel, using 4% methanol in dichlormethane as an eluent, to afford 4-((2,3-dimethyl-l-(4-(4- methylpiperazin- 1 -yl)butyl)- lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dimethoxy- [ 1 , 1 '- biphenyl]-4-yl)benzamide (142 mg, 61%).
1H NMR (400 MHz, DMSO-d6): δ 10.21 (br s, 1H), 7.90 (d, = 8.0 Hz, 2H), 7.41 (d, = 8.4 Hz, 2H), 7.33 (s, 2H), 7.29-7.22 (m, 4H), 7.16 (t, = 9.2 Hz, 2H), 6.94 (br d, = 8.8 Hz, 1H), 4.094.03 (m, 4H), 3.72 (s, 6H), 2.50-2.15 (m, 17H), 1.64- 1.53 (m, 2H), 1.51-1.39 (m, 2H), 1.29-1.20 (m, 2H). LCMS: m/z 663.4 [M+H]+.
Other analogues prepared by this method:
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl-l-(4-(4- methylpiperazin-l-yl)butyl)-lH-indol-5-yl)methyl)benzamide (81%, crude). LCMS: m/z 633.48 [Μ-Η]".
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1-(4-(4- methylpiperazin-l-yl)butyl)-lH-indol-5-yl)methyl)benzamide (63%). LCMS: m/z 644.4 [Μ-Η]". N-(2,6-dimethoxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(4-(4- methylpiperazin-l-yl)butyl)-lH-indol-5-yl)methyl)benzamide (68%). LCMS: m/z 643.4 [Μ-Η]".
N-(4-(lH-imidazol-l-yl)-3,5-dimethoxyphenyl)-4-((2,3-dimethyl-l-(5-(4- methylpiperazin-l-yl)pentyl)-lH-indol-5-yl)methyl)benzamide (66%, crude). LCMS: m/z 649.48 [M+H]+.
N-(3,5-dimethoxy-4-(pyridin-4-yl)phenyl)-4-((2,3 -dimethyl- 1 -(5-(4- methylpiperazin-l-yl)pentyl)-lH-indol-5-yl)methyl)benzamide (61%). LCMS: m/z 660.4 [M+H]+.
N-(2,6-dimethoxy-[l,l'-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(5-(4- methylpiperazin-l-yl)pentyl)-lH-indol-5-yl)methyl)benzamide (70%). LCMS: m/z 659.4 [M+H]+.
4-((2,3-dimethyl-l-(5-(4-methylpiperazin-l-yl)pentyl)-lH-indol-5-yl)methyl)- N-(4'-fluoro-2,6-dimethoxy-[l,r-biphenyl]-4-yl)benzamide (62%). LCMS: m/z 677.4 [M+H]+.
Preparation of Compound 64, 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l- yl)butyl)-lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy-[l,l '-biphenyl]-4- yl)benzamide
To a stirred solution of 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)- lH-indol-5-yl)methyl)-N-(4'-fluoro-2,6-dimethoxy-[l, -biphenyl]-4-yl)benzamide (140 mg, 0.21 mmol) in dichloromethane (6 mL) at 0 °C was added boron tribromide (2.0 mL, 1 M in dichloromethane). The mixture was stirred at this temperature for 10 minutes, then allowed to warm to room temperature and stirred for 12 hours. The reaction mixture was quenched with saturated NaHC03, then extracted with 30% isopropyl alcohol in chloroform. The organic layers were dried over anhydrous Na2S04 and concentrated under reduced pressure to give the crude product. The crude compound was purified by prep-TLC, using 10% methanol in dichloromethane as an eluent, to afford 4-((2,3-dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)-lH-indol-5- yl)methyl)-N-(4'-fluoro-2,6-dihydroxy- [1,1 '-biphenyl] -4-yl)benzamide (Compound 64) as a brown solid (34 mg, 26%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.0 Hz, 2H), 7.39-7.34 (m, 4H), 7.25 (br s, 1H), 7.21 (d, = 8.4 Hz, 1H), 7.06 (t, = 8.8 Hz, 2H), 6.93 (br d, = 8.4 Hz, 1H), 6.85 (s, 2H), 4.19-4.05 (m, 4H), 2.70-2.25 (m, 16H), 2.19 (s, 3H), 1.74- 1.67 (m, 2H), 1.52-1.44 (m, 2H). LCMS: m/z 635.4 [M+H]+.
Other analogues prepared by this method:
Compound 61, N-(3,5-dihydroxy-4-(lH-imidazol- l-yl)phenyl)-4-((2,3- dimethyl-l-(4-(4-methylpiperazin-l-yl)butyl)-lH-indol-5-yl)methyl)benzamide (14%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.0 Hz, 2H), 7.65 (br s, 1H), 7.37 (d, = 8.0 Hz, 2H), 7.26 (br s, 1H), 7.21 (d, = 8.4 Hz, 1H), 7.13 (br s, 1H), 7.05 (br s, 1H), 6.97-6.92 (m, 3H), 4.15-4.11 (m, 4H), 2.64-2.32 (m, 16H), 2.19 (s, 3H), 1.75-1.69 (m, 2H), 1.53-1.46 (m, 2H). LCMS: m/z 607.3 [M+H]+.
Compound 62, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (4-(4-methylpiperazin-l-yl)butyl)-lH-indol-5-yl)methyl)benzamide (23%).
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 6.4 Hz, 2H), 7.81 (d, = 8.4 Hz, 2H), 7.53 (d, = 6.4 Hz, 2H), 7.36 (d, = 8.4 Hz, 2H), 7.26 (br s, 1H), 7.21 (d, = 8.4 Hz, 1H), 6.93 (dd, = 8.4 Hz, 2.0 Hz, 1H), 6.90 (s, 2H), 4.14-4.10 (m, 4H), 2.63- 2.28 (m, 16H), 2.19 (s, 3H), 1.76-1.69 (m, 2H), 1.52- 1.44 (m, 2H). LCMS: m/z
616.23 [M-H]~.
Compound 63, N-(2,6-dihydroxy- [1, 1 '-biphenyl] -4-yl)-4-((2,3 -dimethyl- 1 -(4- (4-methylpiperazin- l-yl)butyl)- lH-indol-5-yl)methyl)benzamide (24%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.0 Hz, 2H), 7.40-7.33 (m, 6H), 7.29-7.20 (m, 3H), 6.93 (br d, = 8.4 Hz, 1H), 6.86 (s, 2H), 4.14-4.09 (m, 4H), 2.72- 2.33 (m, 16H), 2.19 (s, 3H), 1.77-1.69 (m, 2H), 1.52-1.44 (m, 2H). LCMS : m/z 617.3 [M+H]+. Compound 65, N-(3,5-dihydroxy-4-(lH-imidazol- l-yl)phenyl)-4-((2,3- dimethyl-l-(5-(4-methylpiperazin-l-yl)pentyl)-lH-indol-5-yl)methyl)benzamide (15%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.0 Hz, 2H), 7.65 (br s, IH), 7.37 (d, J = 7.6 Hz, 2H), 7.25 (br s, IH), 7.19 (d, J = 8.0 Hz, IH), 7.13 (br s, IH), 7.05 (br s, IH), 6.96 (s, 2H), 6.93 (dd, = 8.4 Hz, 1.6 Hz, IH), 4.12-4.07 (m, 4H), 2.50 (br s, 8H), 2.34-2.30 (m, 8H), 2.19 (s, 3H), 1.72 (quintet, = 7.2 Hz, 2H), 1.47-1.43 (m, 2H), 1.35-1.29 (m, 2H). LCMS: m/z 619.27 [M-H]".
Compound 66, N-(3,5-dihydroxy-4-(pyridin-4-yl)phenyl)-4-((2,3-dimethyl-l- (5-(4-methylpiperazin-l-yl)pentyl)- lH-indol-5-yl)methyl)benzamide (17%).
1H NMR (400 MHz, CD3OD): δ 8.45 (d, = 6.0 Hz, 2H), 7.81 (d, = 8.4 Hz, 2H), 7.53 (d, = 6.0 Hz, 2H), 7.37 (d, = 8.4 Hz, 2H), 7.25 (br s, IH), 7.19 (d, = 8.0 Hz, IH), 6.96-6.88 (m, 3H), 4.12-4.07 (m, 4H), 2.50 (br s, 8H), 2.39-2.30 (m, 8H), 2.19 (s, 3H), 1.74 (quintet, = 7.2 Hz, 2H), 1.51-1.43 (m, 2H), 1.35-1.28 (m, 2H). LCMS: m/z 630.28 [M-H]".
Compound 67, N-(2,6-dihydroxy-[l,r-biphenyl]-4-yl)-4-((2,3-dimethyl-l-(5- (4-methylpiperazin- l-yl)pentyl)-lH-indol-5-yl)methyl)benzamide (28%)
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.4 Hz, 2H), 7.37-7.33 (m, 6H), 7.27-7.23 (m, 2H), 7.19 (d, = 8.4 Hz, IH), 6.93 (dd, = 8.4 Hz, 1.6 Hz, IH), 6.86 (s, 2H), 4.12-4.06 (m, 4H), 2.62-2.27 (m, 16H), 2.19 (s, 3H), 1.73 (quintet, = 7.2 Hz, 2H), 1.50-1.42 (m, 2H), 1.35-1.28 (m, 2H). LCMS: m/z 631.3 [M+H]+.
Compound 68, 4-((2,3-dimethyl- l-(5-(4-methylpiperazin-l-yl)pentyl)- 1H- indol-5-yl)methyl)-N-(4'-fluoro-2,6-dihydroxy-[l, -biphenyl]-4-yl)benzamide (24%).
1H NMR (400 MHz, CD3OD): δ 7.81 (d, = 8.0 Hz, 2H), 7.39-7.35 (m, 4H), 7.25 (br s, IH), 7.19 (d, = 8.4 Hz, IH), 7.06 (t, = 8.8 Hz, 2H), 6.93 (dd, = 8.4 Hz, 1.6 Hz, IH), 6.86 (s, 2H), 4.11-4.07 (m, 4H), 2.54 (br s, 8H), 2.37-2.32 (m, 8H), 2.19 (s, 3H), 1.74 (quintet, = 7.2 Hz, 2H), 1.51-1.43 (m, 2H), 1.36-1.28 (m, 2H). LCMS: m/z 647.26 [M-H]". Scheme 20. Preparation of Compound 69.
Figure imgf000123_0001
Preparation of5-methoxy-2,3-dimethyl-lH-indole
The mixture of (4-methoxyphenyl)hydrazine hydrochloride (10.5 g, 60 mmol) and butan-2-one (10.7 mL, 120 mmol) in acetic acid (200 mL) was heated to reflux for one hour. When cooled, the acetic acid was removed and the residue was dissolved in ethyl acetate (200 mL), washed with saturated NaHC03 and brine, dried over Na2S04 and concentrated in vacuum to give 5-methoxy-2,3-dimethyl- lH-indole as a yellow solid (9.80 g, 94%).
1H NMR (400 MHz, CDC13): δ 7.58 (br s, 1H), 7.14 (d, = 8.8 Hz, 1H), 6.92 (d, J = 1.2 Hz, 1H), 6.76 (dd, = 8.4 Hz, 2.4 Hz, 1H), 3.86 (s, 3H), 2.35 (s, 3H), 2.19 (s, 3H). LCMS: m/z 176.1 [M+H]+.
Preparation of 3-(5-methoxy-2,3-dimethyl-lH-indol-l-yl)-N,N- dimethylpropan-1 -amine To a mixture of NaH (290 mg, 7.24 mmol) in dry DMF (4 mL), was added a solution of 5-methoxy-2,3-dimethyl-lH- indole (317 mg, 1.81 mmol) in dry DMF (1 mL). The mixture was stirred at room temperature for 30 minutes. Then 3-chloro- N,N-dimethylpropan- 1 -amine hydrochloride (572 mg, 3.62 mmol) was added in several portions. After addition, the mixture was stirred at 100 °C for one hour. When cooled, the solution was poured into water and extracted with ethyl acetate. The organic layers were combined, washed with brine, dried over Na2S04 and concentrated in vacuo to give 3-(5-methoxy-2,3-dimethyl-lH-indol-l-yl)-N,N- dimethylpropan-1 -amine as an oil (350 mg, 77%). LCMS: m/z 261.1 [M+H]+.
Preparation of l-( 3-( dimethylamino )propyl)-2,3-dimethyl-lH-indol-5-ol
To a stirring solution of 3-(5-methoxy-2,3-dimethyl-lH-indol-l-yl)-N,N- dimethylpropan-1 -amine (780 mg, 3 mmol) in dichloromethane (10 mL) was added BBr3 (0.5 mL, excess) at 0 °C. The mixture was stirred at room temperature overnight. Ice water (10 mL) was added dropwise, followed by slow addition of saturated Na2C03 (40 mL). The water layer was extracted with ethyl acetate. The organic layers were combined, dried over Na2S04 and concentrated in vacuo to give l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-ol (1.1 g, >100% yield: contaminated with inorganic salt). The crude compound was used in the next step without further purification.
Preparation of 4-( (l-( 3-( dimethylamino )propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoic acid
A mixture of l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-ol (100 mg, 0.407 mmol), Cs2C0 (400 mg, 1.22 mmol) and methyl 4-fluorobenzoate (125 mg, 0.814 mmol) in N-methyl-2-pyrrolidone (4 mL) was stirred at 140 °C overnight. After cooling to room temperature, the mixture was poured into aqueous NaOH (1 M, 5 mL), stirred at room temperature for one hour and then acidified with 1 M HC1 to pH = 4. Water was removed in vacuo. Half of the residue was purified with reverse combi-flash chromatography to give 4-((l-(3-(dimethylamino)propyl)- 2,3-dimethyl-lH-indol-5-yl)oxy)benzoic acid (14 mg, 9%). 1H NMR (400 MHz, CD3OD): δ 7.95 (d, = 7.2 Hz, 2H), 7.40 (br d, J = 3.6 Hz, 1H), 7.15 (br s, 1H), 6.93-6.85 (m, 3H), 4.27 (br s, 2H), 3.17 (br s, 2H), 2.87 (s, 6H), 2.42 (s, 3H), 2.20 (br s, 5H). LCMS: m/z 367.2 [M+H]+.
Preparation of 2-iodo-5-nitrophenol
To a solution of l-iodo-2-methoxy-4-nitrobenzene (1.0 g, 3.6 mmol) in dichloromethane was added BBr3 solution (1 M, 10 mL) at -78 °C. The solution stirred at that temperature for two hours and then warmed to room temperature overnight. Water was added at 0 °C and the mixture was extracted with ethyl acetate, dried over Na2S04 and concentrated in vacuo. The crude product was purified by flash chromatography on silica gel column with ethyl acetate-hexane (1:20), to give 2-iodo-5-nitrophenol as a yellow powder (300 mg, 31%). LCMS: m/z 263.8 [M-H]~.
Preparation of 5-amino-2-iodophenol
To a solution of 2-iodo-5-nitrophenol (300 mg, 1.1 mmol) in ethanol (10 mL), Fe powder (253 mg, 4.5 mmol) was added, followed by concentrated HCl (1 mL). The reaction was stirred at room temperate for one hour. The mixture was filtered and the solution was concentrated to give 5-amino-2-iodophenol (180 mg, 70%). LCMS: m/z 236.0 [M+H]+.
Preparation of Compound 69, 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl- lH-indol-5-yl)oxy)-N-(3-hydroxy-4-iodophenyl)benzamide
A solution of 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoic acid (114 mg, 0.31 mmol) and DIPEA (120 mg, 0.93 mmol) in DMF (4 mL) was stirred at room temperature for 5 minutes, after which HATU (236 mg, 0.62 mmol) was added. The mixture was stirred for 10 minutes prior to the addition of 5-amino-2-iodophenol (73 mg, 0.31 mmol). The resulting mixture was stirred at room temperature overnight. It was then concentrated under vacuum and the residue was partitioned between 1 N HCl and ethyl acetate. The organic layer was separated and concentrated under vacuum. The residue was purified by prep-TLC to provide 4-((l- (3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3-hydroxy-4- iodophenyl)benzamide (Compound 69) as a solid (5 mg, 3%). 1H NMR (400 MHz, CD3OD): δ 7.88 (d, = 8.4 Hz, 2H), 7.61 (d, = 8.8 Hz, 1H), 7.52 (br s, 1H), 7.42 (d, = 8.8 Hz, 1H), 7.16 (br s, 1H), 6.98 (d, = 8.4 Hz, 2H), 6.90-6.86 (m, 2H), 4.29 (t, J = 6.8 Hz, 2H), 3.21-3.16 (m, 2H), 2.92 (s, 6H), 2.43 (s, 3H), 2.21-2.17 (m, 5H).
Scheme 21. Preparation of Compound 70.
Figure imgf000126_0001
Preparation of (2-methoxy-4-nitrophenyl)(methyl)sulfane
A mixture of l-iodo-2-methoxy-4-nitrobenzene (1.9 g, 6.8 mmol), NaSMe (500 mg, 7.1 mmol), Pd2(dba)3 (156 mg), Xantphos (197 mg) and TEA (1.18 mL, 8.51 mmol) in THF (20 mL) was purged with argon and heated to reflux overnight. The reaction mixture was cooled to room temperature and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column (petroleum ether/ethyl acetate, 2: 1, v/v) to give (2-methoxy-4- nitrophenyl)(methyl)sulfane as a yellow solid (400 mg, 30%).
1H NMR (400 MHz, CDC13): δ 7.89 (dd, = 8.8 Hz, 2.4 Hz, 1H), 7.66 (d, = 2.4 Hz, 1H), 7.15 (d, / = 8.8 Hz, 1H), 4.00 (s, 3H), 2.51 (s, 3H).
Preparation of2-methoxy-l -( methylsulfonyl)-4 -nitrobenzene
(2-Methoxy-4-nitrophenyl)(methyl)sulfane (400 mg, 2.0 mmol) was dissolved in methanol (5 mL) and acetonitrile (5 mL). A solution of Oxone (1.9 g, 12 mmol) in water (10 niL) was added and the mixture was stirred at room temperature overnight. The reaction mixture was diluted with water and extracted with ethyl acetate. The combined organic layer was dried over Na2S04 and concentrated under reduced pressure to give 2-methoxy-l-(methylsulfonyl)-4-nitrobenzene as a yellow solid (368 mg, 79%). The product was used in the next step without further purification.
Preparation of 3-methoxy-4-( methylsulfonyl)aniline
A mixture of 2-methoxy-l-(methylsulfonyl)-4-nitrobenzene (368 mg, 1.59 mmol) and SnCl2 (1 g) in ethanol (10 mL) was purged with argon and heated to reflux for one hour. TLC showed that the starting material was consumed completely and a new spot had formed. The reaction mixture was quenched with aqueous NaHC03 and extracted with ethyl acetate. The organic layer was dried over Na2S04 and concentrated under reduced pressure to give 3-methoxy-4-(methylsulfonyl)aniline as a yellow solid (300 mg, 94%). The product was used in the next step without further purification. LCMS: m/z 202.1 [M+H]+.
Preparation of 5-amino-2-(methylsulfonyl)phenol
To a stirring solution of 3-methoxy-4-(methylsulfonyl)aniline (300 mg, 1.5 mmol) in DCM (10 mL) was added BBr3 (1 mL, excess) at 0 °C. The mixture was stirred at room temperature overnight. Ice water (15 mL) was added dropwise, followed by the slow addition of saturated Na2C03 (50 mL). The water layer was extracted with ethyl acetate. The organic layers were combined, dried over Na2S04 and concentrated in vacuo to give 5-amino-2-(methylsulfonyl)phenol (200 mg, 71%). The crude compound was used in the next step without further purification.
Preparation of Compound 70, 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl- lH-indol-5-yl)oxy)-N-(3-hydroxy-4-(methylsulfonyl)phenyl)benzamide
A solution of 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoic acid (114 mg, 0.31 mmol) and DIPEA (120 mg, 0.93 mmol) in DMF (4 mL) was stirred at room temperature for 5 minutes, after which HATU (236 mg, 0.62 mmol) was added. The mixture was stirred for 10 minutes prior to the addition of 5-amino-2-(methylsulfonyl)phenol (58 mg, 0.31 mmol). The resulting mixture was stirred at room temperature overnight. It was then concentrated under vacuum and the residue was partitioned between 1 N HC1 and ethyl acetate. The organic layer was separated and concentrated under vacuum. The residue was purified by prep-TLC to provide 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3- hydroxy-4-(methylsulfonyl)phenyl)benzamide (Compound 70) as a solid (7 mg, 4%).
1H NMR (400 MHz, DMSO-d6): δ 8.09 (d, = 8.4 Hz, 2H), 7.53 (d, = 8.8 Hz, 1H), 7.47 (d, = 8.8 Hz, 1H), 7.22 (br s, 1H), 7.02 (d, = 8.4 Hz, 2H), 6.90 (br d, = 8.4 Hz, 1H), 6.57 (br d, J = 8.8 Hz, 1H), 6.48 (br s, 1H), 6.35 (br s, 2H), 4.16 (t, = 6.8 Hz, 2H), 3.10 (s, 3H), 2.36 (br s, 5H), 2.26 (br s, 6H), 2.16 (s, 3H), 1.86-1.79 (m, 2H).
Scheme 22. Preparation of Compound 71.
Figure imgf000128_0001
Preparation of methyl (E)-3-(2-methoxy-4-nitrophenyl)acrylate
A mixture of l-iodo-2-methoxy-4-nitrobenzene (2.0 g, 7.16 mmol), methyl acrylate (2.0 mL, 22.2 mmol), Pd(OAc)2 (100 mg) and PPh3 (200 mg) in DMF (20 mL) was purged with argon and heated to 70 °C overnight. TLC showed that the starting material was consumed completely and a new spot had formed. The reaction mixture was diluted with ethyl acetate and the catalyst was removed by filtration. The organic layer was washed with brine, dried over Na2S04 and concentrated under reduced pressure. The residue was purified by silica gel column (petroleum ether 100 %) to give methyl (E)-3-(2-methoxy-4-nitrophenyl)acrylate as a yellow solid (500 mg, 31%).
1H NMR (400 MHz, CDC13): δ 7.96 (d, = 16.0 Hz, 1H), 7.84 (dd, = 8.4 Hz, 2.0 Hz, 1H), 7.77 (d, = 2.0 Hz, 1H), 7.63 (d, = 8.4 Hz, 1H), 6.63 (d, = 16.4 Hz, 1H), 4.00 (s, 3H), 3.83 (s, 3H).
Preparation of 7-nitro-2H-chromen-2-one
BBr3 (1 mL, excess) was added dropwise to a solution of methyl (E)-3-(2- methoxy-4-nitrophenyl)acrylate (500 mg, 2.1 mmol) in dichloromethane (5 mL) at 0 °C under argon. The resulting mixture was heated to 50 °C overnight. The cooling reaction mixture was quenched with water and extracted with ethyl acetate. The organic layer was washed with brine, dried over Na2S04 and concentrated under reduced pressure. The residue was purified by silica gel column (petroleum ethenethyl acetate = 2: 1, v/v) to give 7-nitro-2H-chromen-2-one (150 mg, 40%).
1H NMR (400 MHz, CDC13): δ 8.18 (d, = 2.0 Hz, 1H), 8.14 (dd, = 8.4 Hz, 2.4 Hz, 1H), 7.78 (d, = 9.2 Hz, 1H), 7.67 (d, = 8.4 Hz, 1H), 6.62 (d, = 9.6 Hz, 1H).
Preparation of 7 -aminochroman-2-one
A mixture of 7-nitro-2H-chromen-2-one (315 mg, 1.65 mmol), Pd/C (70 mg) and ethyl acetate (25 mL) was stirred at room temperature overnight, under hydrogen pressure of 1.2 MPa. The mixture was then filtered and the filtrate acidified with HCl/ethyl acetate. The resulting precipitate was collected by filtration and dried to give 7-aminochroman-2-one (105 mg, 40%).
1H NMR (400 MHz, DMSO-d6): δ 9.80 (br s, 2H), 7.35 (d, = 8.4 Hz, 1H), 7.04 (dd, = 8.0 Hz, 2.0 Hz, 1H), 6.98 (d, = 2.0 Hz, 1H), 2.98 (t, = 7.2 Hz, 2H), 2.79 (t, 7 = 7.2 Hz, 2H).
Preparation of 4-( (l-( 3-( dimethylamino )propyl)-2,3-dimethyl-lH-indol-5- yl )oxy )-N-(2 -oxochroman-7-yl )benzamide A solution of 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoic acid (114 mg, 0.31 mmol) and DIPEA (120 mg, 0.93 mmol) in DMF (4 mL) was stirred at room temperature for 5 minutes, after which HATU (236 mg, 0.62 mmol) was added. The mixture was stirred for 10 minutes prior to the addition of 5-amino-2-(methylsulfonyl)phenol (51 mg, 0.31 mmol). The resulting mixture was stirred at room temperature overnight. It was then concentrated under vacuum and the residue was partitioned between 1 N HC1 and ethyl acetate. The organic layer was separated and concentrated under vacuum. The residue was purified by prep-TLC to provide 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(2- oxochroman-7-yl)benzamide as a solid (19 mg, 12%).
Preparation of Compound 71, 3-(4-(4-((l-(3-(dimethylamino)propyl)-2,3- dimethyl-lH-indol-5-yl)oxy)benzamido)-2-hydroxyphenyl)propanoic acid
To a solution of 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)-N-(2-oxochroman-7-yl)benzamide (19 mg, 0.04 mmol) in THF (5 mL) was added aqueous LiOH (4 N, 5 mL). The mixture was stirred at room temperature for three hours, then acidified to pH = 4 with 1 N HC1 aqueous solution and extracted with ethyl acetate. The organic layers were combined, dried over anhydrous Na2S04, filtered and concentrated under reduced pressure to give 3-(4-(4-((l-(3- (dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)benzamido)-2- hydroxyphenyl)propanoic acid (Compound 71) as a solid (5 mg, 26%).
1H NMR (400 MHz, DMSO-d6): δ 9.89 (br s, 1H), 7.91 (d, = 8.4 Hz, 2H), 7.43 (d, J = 8.8 Hz, 1H), 7.39 (br s, 1H), 7.14 (d, J = 1.6 Hz, 1H), 7.02-6.95 (m, 4H), 6.86 (dd, = 8.8 Hz, 1.6 Hz, 1H), 4.14 (t, = 6.8 Hz, 2H), 2.73-2.68 (m, 2H), 2.46- 2.42 (m, 2H), 2.35 (br s, 5H), 2.24 (s, 6H), 2.14 (s, 3H), 1.85-1.79 (m, 2H). LCMS: m/z 530.3 [M+H]+.
Scheme 23. Preparation of Compound 72.
Figure imgf000131_0001
Preparation of 2-methoxy-4-nitrobenzonitrile
To a mixture of CuCN (322 mg, 3.6 mmol) and L-proline (415 mg, 3 mmol), in DMF (15 mL) under argon, l-iodo-2-methoxy-4-nitrobenzene (837 mg, 3 mmol) was added at room temperature. The mixture was stirred at 80 °C for 45 hours. The resulting suspension was cooled to room temperature, diluted with ethyl acetate (15 mL), and washed with water. The organic phase was dried over Na2S04 and concentrated. The residue was purified by flash chromatography on silica with ethyl acetate-hexane to yield 2-methoxy-4-nitrobenzonitrile (463 mg, 87%).
Preparation of 5-(2-methoxy-4-nitrophenyl)-2H-tetrazole
A mixture of 2-methoxy-4-nitrobenzonitrile (510 mg, 2.76 mmol), NaN3 (719 mg, 11.06 mmol) and ZnBr2 (2.50 g, 11.06 mmol) in water (5 mL) was heated at 160 °C for 30 minutes under microwave conditions. Chromatographic purification afforded 5-(2-methoxy-4-nitrophenyl)-2H-tetrazole (450 mg, 75%).
Preparation of 5-nitro-2-(2H-tetrazol-5-yl)phenol
To a solution of 5-(2-methoxy-4-nitrophenyl)-2H-tetrazole (200 mg, 0.90 mmol) in dichloromethane (10 mL) was added BBr3/CH2Cl2 (0.5 mL, excess) dropwise at -78 °C with stirring. The resulting mixture was stirred at room temperature for one hour and then worked up with ice-water and extracted with dichloromethane. The organic layer was dried over anhydrous Na2S04, concentrated in vacuo and purified by prep-TLC to give 5-nitro-2-(2H-tetrazol-5-yl)phenol (120 mg, 64%).
Preparation of 5-amino-2-(2H-tetrazol-5-yl)phenol
A mixture of 5-nitro-2-(2H-tetrazol-5-yl)phenol (120 mg, 0.58 mmol) and Pd/C (30 mg) in MeOH (5 mL) was charged with H2 and stirred at room temperature overnight. After chromatographic purification, 5-amino-2-(2H-tetrazol-5-yl)phenol was obtained (87 mg, 84%).
Preparation of Compound 72, 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl- lH-indol-5-yl)oxy)-N-(3-hydroxy-4-(2H-tetrazol-5-yl)phenyl)benzamide
A solution of 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5- yl)oxy)benzoic acid (114 mg, 0.31 mmol) and DIPEA (120 mg, 0.93 mmol) in DMF (4 mL) was stirred at room temperature for 5 minutes, after which HATU (236 mg, 0.62 mmol) was added. The mixture was stirred for 10 minutes prior to the addition of 5-amino-2-iodophenol (55 mg, 0.31 mmol). The resulting mixture was stirred at room temperature overnight. It was then concentrated under vacuum and the residue was partitioned between 1 N HCl and ethyl acetate. The organic layer was separated and concentrated under vacuum. The residue was purified by prep-TLC to provide 4-((l-(3-(dimethylamino)propyl)-2,3-dimethyl-lH-indol-5-yl)oxy)-N-(3-hydroxy-4- (2H-tetrazol-5-yl)phenyl)benzamide (Compound 72) as a solid (6 mg, 4%).
1H NMR (400 MHz, CD3OD): δ 8.04 (d, = 8.4 Hz, 1H), 7.93 (d, = 8.4 Hz, 2H), 7.85 (br s, 1H), 7.43-7.40 (m, 2H), 7.18 (br s, 1H), 7.01 (d, = 8.8 Hz, 2H), 6.94-6.88 (m, 2H), 4.28 (t, J = 6.8 Hz, 2H), 3.21-3.16 (m, 2H), 2.89 (s, 6H), 2.44 (s, 3H), 2.22-2.14 (m, 5H). LCMS: m/z 526.4 [M+H]+.
Example 2 - Activity of anti-tropomyosin compounds as monotherapy
In silico modelling has identified binding sites on tropomyosin Tpm3.1 yielding the series of tropomyosin inhibitors the subject of the present invention. Inhibition of Tpm3.1 in tumour cells results in disruption of the actin cytoskeleton and ultimately cell death. The ability of compounds 50 and 51 to disrupt the total actin cytoskeleton (Figure 1) and to specifically target Tpm3.1 -containing actin filaments (Figure 2) was assessed in vitro using the microfilament disruption assay.
Briefly, SK-N-SH neuroblastoma cells were seeded at 30 x 10 cells/well in a in a volume of 1000 μΐ^ with complete media onto a 12 well plate containing 19 mm coverslips and left to plate down for 24 hours prior to treatment. Cells were then treated with 0, 2.5 and 5 μΜ of the test compounds. 24 hours post treatment, cells were fixed with 4% paraformaldehyde (PBS) and stained with either 488-Atto- Phallodin and DAPI to visualize the actin filament bundles and the nucleus, or with j9d primary antibody (MAb culture s/n clone 2G10.2, 1:50) followed by 488- conjugated secondary antibody (Goat anti mouse 488, 1: 1000) and DAPI to visualize the Tpm3.1 -containing filament bundles and the nucleus, respectively. Single plane images were obtained on the Perkin Elmer Opera confocal microscope using a 20x objective. Twelve fields of view per condition were imaged. Images were then exported and changes in the organization and numbers of actin filaments within the cell were quantitated using a linear feature detection algorithm developed by the Commonwealth Scientific and Industrial Research Organisation (CSIRO). This algorithm detects the "ridge lines" or "peaks" in local pixel intensity in the cell image. It is these "ridge lines" that correspond to actin filament bundles and allow us to quantitate the number of filaments per cell.
Data demonstrate that compounds 50 and 51 disrupt both the total actin cytoskeleton and Tpm3.1 -containing actin filaments in a dose-dependent manner.
The ability of compounds 1-12, 14-20, 26-28, 33-36, 38-40, and 42-68 to inhibit the proliferation of cancer cells representative of neuroblastoma, melanoma, prostate cancer, colorectal cancer, non-small cell lung carcinoma, and triple negative breast cancer was assessed (see Table 1). These studies were conducted by contract research (GVK-BIO). Briefly, a pre-determined number of cells as calculated from cell growth assays for each of the cell lines employed were seeded into their respective culture medium (using ATCC culture parameters - htt : //www . atcc . org ) and cultured for 24 h at 37 °C and 5% C02 in 96-well culture plates. Once attached, each cell line was then exposed to various concentrations of each respective analogue (30, 10, 3, 1, 0.3 and 0.1 μΜ), cultured for a further 72 h and exposed to cell-titre luminescent reagent (100 μΐ/ννεΐΐ) for a further 30 min. Luminescence was captured using an En Vision multilabel reader and the data for each analogue concentration compared against no treatment control. Cell viability was normalized to control (vehicle alone) and dose-response curves, and half maximal effective concentration (EC50) values were determined using Graph Pad Prism 6 (nonlinear regression sigmoidal dose-response variable slope).
Table 1. Anti-proliferative activity of compounds of the invention against a range of somatic cancer cells
Ι050 / μΜ
NeuroblasLung toma Melanoma Prostate Colorectal (NSLC) Breast
Compound MDA-MB- ID SK-N-SH SK-Mel-28 DU145 PC3 CaCo2 A549 231
1 >30 >30 >30 >30 >30 >30 >30
2 7.1 5.5 >30 >30 9.2 >30 8.9
3 7.8 9.1 >30 8.9 7.4 >30 10.3
4 2.4 3.8 4.2 4.0 2.6 3.8 2.8
5 >30 5.1 >30 >30 >30 >30 23.3
6 3.3 1.3 3.6 1.8 1.2 2.8 1.2
7 4.6 1.4 3.4 2.3 1.2 2.6 1.6
8 1.1 1.5 3.6 1.9 2.2 3.0 1.4
9 >30 >30 >30 >30 >30 3.7 >30
10 6.3 19.4 >30 30 17.5 3.7 9.9
11 5.0 4.6 3.8 5.1 10.2 3.7 3.6
12 5.0 4.7 4.0 8.2 9.7 3.7 4.0
14 1.4 0.9 3.2 1.9 2.1 2.9 1.7
15 3.8 3.0 3.5 3.6 3.5 2.6 3.4
16 0.4 0.3 0.3 0.3 0.4 0.3 0.2
17 >30 >30 >30 >30 >30 10.2 >30
18 7.7 4.9 10.5 7.8 3.8 10.2 3.9
19 6.3 4.9 10.7 9.0 5.4 9.3 5.0
20 5.4 3.8 3.6 3.9 3.3 3.6 3.5
26 9.5 1.4 3.1 1.7 1.3 2.1 1.3
27 6.0 1.2 2.2 1.3 1.2 1.4 1.1
28 2.0 3.8 10.7 4.6 3.8 11.2 3.6
33 >30 >30 >30 >30 >30 >30 >30
34 3.5 4.5 9.2 5.4 10.5 18.8 5.0
35 4.6 2.8 4.4 6.4 2.1 3.7 2.5
36 1.1 4.2 3.7 3.2 3.5 3.5 4.1
38 11.7 1.3 2.1 1.9 1.2 1.4 1.3
39 >30 1.3 2.3 1.5 1.3 1.2 1.3
Figure imgf000135_0001
The anti-proliferative activity of compounds 69-72 against SHEP neuroblastoma and SK-Mel-28 melanoma cells was also assessed. SHEP and SK-Mel-28 cell lines were maintained as monolayers in Dulbeccos Modified Eagles medium (DMEM-Invitrogen) supplemented with 10% fetal bovine serum (FBS) and grown at 37 °C in a humidified atmosphere with 5% C02. For the cytotoxicity assay, 2 x 10J cells were plated into 96 well plates and treated with serial dilutions of drug (1:2 starting concentration of 200 μΜ) and viability measured after 48 hours using a standard MTS assay. Cell viability was normalised to control (vehicle alone) and dose response curves and EC50 values (Table 2) were determined using Graph Pad Prism 5 (non linear regression sigmoidal dose response-variable slope). Table 2. Anti-proliferative activity of Compounds 69-72 against neuroblastoma and melanoma cells
Figure imgf000136_0001
Example 3 - Activity of compounds of formula (I) in combination with vincristine and paclitaxel
A study was performed to determine synergistic drug interactions between the cytotoxic agents vincristine and paclitaxel in combination with compounds 6, 8, 14, 16, 23, 24, 27, 31, 32, 38, 39, 47, 48, 51, 55, 58, 60, 63, 67 and 68 against A375 (malignant melanoma), A549 (lung carcinoma) and DU145 (prostate carcinoma) cell lines.
3.1 Materials and methods Compounds
Stock solutions (10 mM) were prepared by dissolving compounds in 100% sterile DMSO. Vincristine and paclitaxel were further diluted to 0.1 mM for experiments. Small aliquots (20 μΐ) were prepared and stored at -30 °C under minimum light. Stock solutions were defrosted once only for each experiment.
Cell lines
All cell lines (A375, A549 and DU145) were tested for mycoplasma contamination and tested negative.
Cell expansion and culture conditions
The A549 and A375 cells were cultured in DMEM cell culture medium supplemented with 10% (v/v) FBS and the DU145 cells were cultured in RPMI cell culture medium supplemented with 10% (v/v) FBS. All cell lines were cultured in the absence of antibiotics for no more than 10 passages. The cells were harvested at -80% confluence by trypsinisation (passage +111 (A549), passage +20 (A375) and passage 8 (DU145)). Cells were then washed, counted and plated in 384-well plates.
In vitro growth inhibition assay
In vitro growth inhibition assays were performed as technical triplicate in two independent experiments and cell viability was assessed by alamar blue assay readout. Briefly, cells in logarithmic phase growth were seeded in 384-well plates in a 50μΙ^Λνε11 assay volume. Cells were seeded at 250 cells/well for A549, 400 cells/well for A375 and 500 cells/well for DU145 cells using a Multidrop 384 (Thermo Scientific) and allowed to adhere at 37 °C with 5% C02, in a humidified atmosphere (LiCONiC incubator). After 24 hours incubation, test compounds, positive control (20 μΜ Thonzonium bromide, 100% kill) and vehicle only (0.4% DMSO) were added into wells of each assay plate using a Tecan HP D300 Digital Dispenser in technical triplicate under minimum light. Following 72 hours drug exposure, metabolic activity was detected by addition of 10% (v/v) alamar blue reagent to each assay well using a Multidrop Combi (Thermo Scientific) and determined by measurement of fluorescence intensity (excitation 555 nm, emission 585 nm) using an EnSpire (Perkin Elmer) plate reader. Readings were performed at 0 hour (background) and after 6 hours alamar blue incubation at 37 °C.
IC50 determination
IC50 concentrations of the compounds together with vincristine and paclitaxel as single agents against all cell lines were determined for a 72 h drug exposure using the growth inhibition assay with alamar blue readout in triplicate in two independent experiments. Each drug was tested as a 10-point concentration series with two-fold serial dilutions of each drug. The concentration series for the compounds for all cell lines tested was 20, 10, 5, 2.5, 1.25, 0.625, 0.3125, 0.156, 0.078 and 0.039 μΜ for the two independent experiments. The concentration series for vincristine and paclitaxel was 100, 50, 25, 12.5, 6.25, 3.125, 1.5625, 0.781, 0.391 and 0.195 nM. Thonzonium bromide (20 μΜ) and 0.4% DMSO were used as positive and vehicle controls, respectively. The IC50 value was calculated by derivation of the best-fit line using a sigmoidal dose response model (Activity Base Software Suite, IDBS). Drug combination screen
Each drug combination was tested against A375, A549 and DU145 cells using a 6x6 dose matrix with 2-fold dilution steps based on the calculated IC50 values using the growth inhibition assay with 72h drug exposure in a 384-well format. The two drugs in each combination matrix were dispensed using a Tecan HP D300 Digital Dispenser. The layout of each screening plate is shown in Figure 3. Assays were performed in triplicate for each drug combination (ATM/vincristine and ATM/paclitaxel combinations). Synergistic drug interactions were calculated from the averaged cell viability data using the methodology described below.
3.2 Data analyses
Activity Base software suite (IDBS, version 8.3.0.175) was used for data analysis. Cell survival is expressed as a percentage of viable cells. IC50 concentrations were calculated by derivation of the best-fit line using a sigmoidal dose response model with technical triplicate data points.
Methodology for calculation of synergistic drug interactions
Synergistic drug interactions were determined for each pairwise drug combination by applying the Bliss-independence model that assumes independence of drug mechanisms. Prior to the analysis, percent cell viability (CV) for combination data were converted to fractional cell growth inhibition (GI) by the formula
GI = 1-(CV/100)
The Bliss additivism model was used to calculate a predicted combined response C to two single agents with responses A and B (using GI values), as follows:
C = A + B - (A x B)
Experimentally observed values in excess of the predicted additive value for each combination demonstrated a synergistic effect while values that were lower than the predicted additive value demonstrated antagonism of inhibition of cell viability relative to single agent alone. The final score for any combination was reported as the difference between the value predicted by the additive model and the experimentally observed value, normalised to 100. Using GI values the maximum possible synergy score is 100, with 0 indicating no synergy and negative indicating possible antagonism. Scores were also reported as a "Max synergy" score, which is the highest individual synergy score for each combination matrix, and a "Total synergy" score, which was obtained by summation of each individual synergy score for the matrix.
3.3 Results IC50 determination
To enable generation of the 6x6 dose matrices for each drug combination, the IC50 concentrations of the compounds, vincristine and paclitaxel as single agents were determined against A375, A549 and DU145 cells using the growth inhibition assay with alamar blue cell viability readout. The data is listed as IC50 mean from technical triplicates for two independent experiments in Table 3. The averaged IC50 concentrations for the single agents from both runs were then used to generate the 6x6 dose matrices in the synergy experiment.
Table 3. The ICgn values of the compounds, vincristine and paclitaxel in A375, A549 and DU145 cells assessed by in vitro growth inhibition assay*
Figure imgf000139_0001
Figure imgf000140_0001
*Dose response curves of ATM-3000 compounds, vincristine and paclitaxel with 10 dose points at 72 hours post-treatment were generated, allowing for the calculation of ICsn values. The averaged ICsn values from two independent experiments were used to generate 6x6 dose matrices in the synergy experiment. Drug combination screen
The compounds were tested in combination with vincristine and paclitaxel as a 6x6 dose matrix in triplicate for each cell line. Synergy scores for each drug combination were calculated by the Bliss-independence model. The max synergy and total synergy scores for each combination in all cell lines tested are listed in Tables 4- 6 below.
Table 4. Synergy scores for each drug combination matrix in A375 cellss
Figure imgf000141_0001
23 Vincristine 55.82 526.03
16 Vincristine 49.28 499.59
14 Vincristine 39.45 312.10
8 Vincristine 61 .53 500.66
6 Vincristine 42.53 316.03
68 Paclitaxel 26.80 319.39
67 Paclitaxel 45.12 507.55
63 Paclitaxel 29.07 377.42
60 Paclitaxel 35.66 351 .68
58 Paclitaxel 46.1 5 368.55
55 Paclitaxel 44.99 567.84
51 Paclitaxel 26.96 316.38
48 Paclitaxel 34.54 323.08
47 Paclitaxel 31 .46 297.82
39 Paclitaxel 25.35 259.27
38 Paclitaxel 36.38 232.91
32 Paclitaxel 29.40 246.34
31 Paclitaxel 23.91 189.84
27 Paclitaxel 28.71 237.09
24 Paclitaxel 34.68 266.00
23 Paclitaxel 28.27 242.88
16 Paclitaxel 39.93 361 .50 14 Paclitaxel 33.27 249.51
8 Paclitaxel 30.88 271 .50
6 Paclitaxel 49.55 412.70
* Max and Total synergy scores were determined for each drug combination matrix. Max synergy is the highest individual synergy score for each combination matrix; Total synergy is the summation of each individual synergy score for the matrix.
Table 5. Synergy scores for each drug combination matrix in A549 cells
68 Vincristine 21 .25 136.36
67 Vincristine 20.97 125.60
63 Vincristine 28.63 185.53
60 Vincristine 39.52 234.89
58 Vincristine 36.26 142.30
55 Vincristine 16.07 1 16.79
51 Vincristine 16.17 56.58
48 Vincristine 32.40 134.29
47 Vincristine 26.14 126.30
39 Vincristine 26.42 1 10.88
38 Vincristine 24.69 95.23
32 Vincristine 17.04 1 12.01
31 Vincristine 20.1 1 1 10.63
27 Vincristine 27.48 140.70
24 Vincristine 24.1 1 184.55 23 Vincristine 21 .62 140.85
16 Vincristine 45.76 258.19
14 Vincristine 41 .28 205.40
8 Vincristine 26.73 190.44
6 Vincristine 49.75 248.58
68 Paclitaxel 7.63 -28.51
67 Paclitaxel 10.1 7 30.34
63 Paclitaxel 9.33 -15.29
60 Paclitaxel 12.05 48.19
58 Paclitaxel 7.67 0.48
55 Paclitaxel 1 1 .1 7 49.21
51 Paclitaxel 18.39 57.15
48 Paclitaxel 9.19 49.62
47 Paclitaxel 7.01 43.85
39 Paclitaxel 10.78 27.42
38 Paclitaxel 6.07 -4.87
32 Paclitaxel 10.07 3.46
31 Paclitaxel 5.98 4.56
27 Paclitaxel 8.44 15.40
24 Paclitaxel 1 .71 ~39.59
23 Paclitaxel 4.04 -17.61
16 Paclitaxel 5.62 13.63 14 Paclitaxel 12.59 9.94
8 Paclitaxel 10.72 5.69
6 Paclitaxel 20.29 83.32
*Max and Total synergy scores were determined for each drug combination matrix. Max synergy is the highest individual synergy score for each combination matrix; Total synergy is the summation of each individual synergy score for the matrix.
Table 6. Synergy scores for each drug combination matrix in DU145 cells'
68 Vincristine 70.83 419.75
69 Vincristine 55.40 200.97
63 Vincristine 63.09 241 .08
60 Vincristine 81 .38 380.32
58 Vincristine 82.81 435.74
55 Vincristine 35.06 103.17
51 Vincristine 54.46 223.32
48 Vincristine 48.06 254.64
47 Vincristine 45.89 235.17
39 Vincristine 57.80 345.06
38 Vincristine 54.13 294.28
32 Vincristine 52.60 252.08
31 Vincristine 40.1 2 214.12
27 Vincristine 45.31 254.40
24 Vincristine 77.05 528.65 23 Vincristine 83.1 6 506.43
16 Vincristine 86.43 376.42
14 Vincristine 83.34 422.03
8 Vincristine 85.57 480.99
6 Vincristine 83.25 480.45
68 Paclitaxel 16.34 73.62
67 Paclitaxel 15.44 73.25
63 Paclitaxel 12.72 56.36
60 Paclitaxel 1 1 .48 65.72
58 Paclitaxel 13.03 82.84
55 Paclitaxel 14.67 52.02
51 Paclitaxel 22.76 121 .27
48 Paclitaxel 5.21 33.28
47 Paclitaxel 5.34 26.54
39 Paclitaxel 10.91 17.25
38 Paclitaxel 6.25 8.84
32 Paclitaxel 5.70 4.83
31 Paclitaxel 10.97 42.79
27 Paclitaxel 7.78 28.32
24 Paclitaxel 19.19 88.85
23 Paclitaxel 19.13 75.57
16 Paclitaxel 26.44 106.14 14 Paclitaxel 25.85 1 10.16 8 Paclitaxel 16.56 104.85
6 Paclitaxel 12.62 67.09
* Max and Total synergy scores were determined for each drug combination matrix. Max synergy is the highest individual synergy score for each combination matrix; Total synergy is the summation of each individual synergy score for the matrix.
The combination of vincristine and compounds of the formula (I) yielded medium to high level synergistic enhancement of cell death for A375, A549 and DU145 cells, with the highest degree of synergy observed for DU145 cells. In the case of compounds of formula (I) in combination with paclitaxel a higher synergy score was observed in A375 cells compared to the other two cell lines tested.
The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgement or admission or any form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.
Those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention includes all such variations and modifications. The invention also includes all of the steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of any two or more of said steps, features, compositions and compounds.

Claims

1. A compound of the formula (I),
Figure imgf000148_0001
(I) or a pharmaceutically acceptable salt, hydrate, derivative, solvate or prodrug thereof, wherein:
1 2
R and R" are independently selected from the group consisting of hydrogen and Ci- C6 alkyl;
3 8 10
RJ is NR°R1U or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6 and wherein the ring is optionally substituted with R ;
R4 is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, O, S, or NR6;
R5 is
Figure imgf000148_0002
each Y is independently OH or Ci-C6 alkoxy and X is a 5- or 6-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with between 1 and 3 substituents selected from the group consisting of: halo and Ci-C6 alkyl, or
R is Q wherein Z is OH or Ci-C6 alkoxy and Q is selected from the group consisting of: halo, -S02Ci-C6 alkyl, -(CH2)0-5COOH, -(CH2)0-5COOCi-C6 alkyl or a 5- or 6-membered carbocyclic ring wherein between 1 and 4 carbons may optionally be replaced by N or NR6;
R6 is selected from the group consisting of: H and Ci-C6 alkyl;
R is selected from the group consisting of: H, halo, Ci-C6 alkyl, Ci-C6alkoxy, CN, CF3 and OCF3;
R 8° and R 110U are independently selected from the group consisting of: H and Ci-C6 alkyl;
X1 is an alkanediyl group having between 1 and 20 carbon atoms; and
X 2" and 3
X3 are independently selected from the group consisting of: -0-, -NH-, -NCi- C6alkyl-, -C(O)-, -C(0)NH-, -NHC(O)- or an alkanediyl group having between 1 and 20 carbon atoms wherein between 1 and 3 hydrogen atoms may optionally be replaced with R .
2. The compound of claim 1, wherein R 1 and R 2 are independently selected from the group consisting of: H and Ci-C3 alkyl.
3. The compound of claim 2, wherein R 1 and R 2 are both methyl.
4. The compound of any one of claims 1 to 3, wherein R 3 is NR 8 R 10 or a 4- to 7-membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6 and wherein the ring is optionally substituted with R7.
5. The compound of claim 4, wherein R 3 is NR 8 R 10 or a 5- or 6- membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N or NR6.
6. The compound of claim 5, wherein R is N(Me)2, NH2 or a 5- or 6- membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N or NR6.
7. The compound of claim 6, wherein R is N(Me)2 or a 6-membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
8. The compound of claim 4, wherein R is N(Me)2 or a saturated 6- membered carbocyclic ring wherein 1 or 2 carbons may optionally be replaced by N, NH or NMe.
9. The compound of any one of claims 1 to 3, wherein R is selected from the group consisting of: N(Me)2, NH2 and
Figure imgf000150_0001
10. The compound of any one of claims 1 to 3, wherein R is selected from
Figure imgf000150_0002
the group consisting of: N(Me)2 and
11. The compound of any one of claims 1 to 10, wherein R4 is a 5- or 6- membered carbocyclic ring wherein between 1 and 3 carbons may optionally be replaced by N, NR6, O or S.
12. The compound of claim 11, wherein R4 is phenylene or a 5- or 6- membered heteroarylene ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S.
13. The compound of claim 12, wherein R4 is phenylene or a 5- or 6- membered heteroarylene ring having between 1 and 3 N heteroatoms.
14. The compound of claim 13, wherein R4 is phenylene or pyridylene.
15. The compound of claim 14, wherein R4 is phenylene.
16. The compound of claim 15, wherein R4 is para-phenylene or meta- phenylene.
The compound of any one of claims 1 to 16, wherein R
Figure imgf000151_0001
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having between 1 and 3 heteroatoms selected from the group consisting of: N, O and S, and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 substituents selected from the group consisting of: halo, methyl, ethyl, propyl and isopropyl, or
R5 is
Figure imgf000151_0002
Z is OH or OMe and Q is selected from the group consisting of: halo, -S02Me, -(CH2)0-5COOH, -(CH2)0-5COOCi-C6alkyl, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
18. The compound of claim 17, wherein R is
Figure imgf000151_0003
wherein Y is OH or OMe and X is phenyl or a 5- or 6- membered heteroaryl ring having between 1 and 3 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with 1 or 2 halo substituents, or
R5 is
Figure imgf000151_0004
Z is OH or OMe and Q is selected from the group consisting of: halo, -S02Me, -(CH2)i_sCOOH, phenyl and a 5- or 6-membered heteroaryl ring having between 1 and 4 N heteroatoms.
19. The compound of claim 18, wherein R is
Figure imgf000151_0005
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a halo substituent, or
R5 is
Figure imgf000152_0001
wherein Z is OH or OMe and Q is selected from the group consisting of: I, -S02Me, -(CH2)i-3COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms.
20. The compound of claim 19, wherein R5 is
Figure imgf000152_0002
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or
R5 is
Figure imgf000152_0003
Z is OH and Q is selected from the group consisting of: I, -S02Me, -(CH2)2COOH, or a 5-membered heteroaryl ring having between 1 and 4 N heteroatoms.
21. The compound of claim 20, wherein R5 is
Figure imgf000152_0004
wherein Y is OH or OMe and X is phenyl or a 5- or 6-membered heteroaryl ring having 1 or 2 N heteroatoms, and wherein the phenyl and the heteroaryl ring may optionally be substituted with a fluoro substituent, or R is K Q wherein Z is OH and Q is selected from the group consisting of: I, -S02Me, -(CH2)2COOH and tetrazolyl.
22. The compound of claim 21, wherein X is selected from the group consisting of:
Figure imgf000153_0001
23. The compound of claim 21, wherein X is selected from the group consisting of:
Figure imgf000153_0002
nd of any one of claims 1 to 23, wherein R5
Figure imgf000153_0003
The compound of any one of claims 1 to 23, wherein R5
Figure imgf000153_0004
26. The compound of any one of claims 1 to 25, wherein X1 is an alkanediyl group having between 1 and 10 carbon atoms.
27. The compound of claim 26, wherein X1 is an alkanediyl group having between 1 and 6 carbon atoms.
28. The compound of claim 27, wherein X1 is -(CH2)i_6- or -CH2-CH(Me)-
(CH2)-.
29. The compound of any one of claims 1 to 28, wherein X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, - NHC(O)- and an alkanediyl group having between 1 and 10 carbon atoms.
30. The compound of claim 29, wherein X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -NMe, -C(0)NH-, -NHC(O)- and an alkanediyl group having between 1 and 6 carbon atoms.
31. The compound of claim 30, wherein X 2 and X 3 are independently selected from the group consisting of: -0-, -NH-, -C(0)NH-, -NHC(O)- and CH2.
32. The compound of any one of claims 1 to 28, wherein X is selected from the group consisting of: -0-, -NH-and -CH2-.
33. The compound of any one of claims 1 to 28, wherein X is -C(0)NH- or -NHC(O)-.
34. A compound according to claim 1, wherein the compound is selected from the group consisting of:
Figure imgf000155_0001
Figure imgf000155_0002
Figure imgf000155_0003
Figure imgf000155_0004
Figure imgf000156_0001
Figure imgf000156_0002
Figure imgf000156_0003
Figure imgf000156_0004
15 16
Figure imgf000157_0001
Figure imgf000158_0001
Figure imgf000158_0002
Figure imgf000158_0003
Figure imgf000158_0004
31 32
Figure imgf000159_0001
Figure imgf000160_0001
Figure imgf000160_0002
Figure imgf000160_0003
Figure imgf000160_0004
47 48
Figure imgf000161_0001
Figure imgf000161_0002
Figure imgf000161_0003
Figure imgf000161_0004
55 56
Figure imgf000162_0001
Figure imgf000162_0002
Figure imgf000162_0003
Figure imgf000163_0001
Figure imgf000163_0002
Figure imgf000163_0003
Figure imgf000163_0004
35. A compound according to claim 34, wherein the compound is selected from compounds 6, 8, 16, 23, 24, 47, 48, 51, 60 and 68.
36. A pharmaceutical composition comprising a compound of any one of claims 1 to 35 together with a pharmaceutically acceptable carrier, diluent or excipient.
37. A method for the treatment of a proliferative disease in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound according to any one of claims 1 to 35, or a pharmaceutical composition according to claim 36.
38. The method of claim 37, wherein the proliferative disease is cancer.
39. The method of claim 38, wherein the cancer is breast cancer, lung cancer, prostate cancer, colon cancer, melanoma or neuroblastoma.
40. The method of claim 38 or claim 39, wherein the cancer has recurred.
41. Use of a compound according to any one of claims 1 to 35 in the manufacture of a medicament for the treatment of a proliferative disease.
42. A method for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence, the method comprising administration to the subject of an effective amount of a compound according to any one of claims 1 to 35, or a pharmaceutical composition according to claim 36.
43. Use of a compound of formula (I) according to any one of claims 1 to 35 in the manufacture of a medicament for reducing incidences of, or risk of, cancer recurrence in a subject deemed to be at risk of cancer recurrence.
44. The method of claim 42, wherein the subject is a subject who is in cancer remission.
45. A combination comprising:
(i) a compound of the general formula (I) according to any one of claims
1 to 34,
and,
(ii) vincristine or paclitaxel.
46. A pharmaceutical composition comprising:
(i) a compound of the general formula (I) according to any one of claims
1 to 34,
(ii) vincristine or paclitaxel, and
(iii) a pharmaceutically acceptable carrier, diluent or excipient.
47. A kit comprising: (i) a pharmaceutical composition of a compound of the general formula (I) according to any one of claims 1 to 34,
and
(ii) a pharmaceutical composition of vincristine or paclitaxel, wherein the compositions are intended for simultaneous, concurrent, separate or sequential use.
48. A method for the treatment of cancer in a subject in need thereof, the method comprising administration to the subject of a therapeutically effective amount of a compound of formula (I) according to any one of claims 1 to 34, and a therapeutically effective amount of vincristine or paclitaxel.
49. The method of claim 48, wherein the cancer is melanoma, lung cancer or prostate cancer.
50 The combination of claim 45, the composition of claim 46, or the kit of claim 47, wherein the compound of the general formula (I) is selected from one or more of compounds 6, 8, 14, 16, 23, 24, 27, 31, 32, 38, 39, 47, 48, 51, 55, 58, 60, 63, 67 and 68.
51. The method of claim 48 or claim 49, wherein the compound of the general formula (I) is selected from one or more of compounds 6, 8, 14, 16, 23, 24, 27, 31, 32, 38, 39, 47, 48, 51, 55, 58, 60, 63, 67 and 68.
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