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WO2004041812A1 - Composes et methodes therapeutiques - Google Patents

Composes et methodes therapeutiques Download PDF

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Publication number
WO2004041812A1
WO2004041812A1 PCT/AU2003/001463 AU0301463W WO2004041812A1 WO 2004041812 A1 WO2004041812 A1 WO 2004041812A1 AU 0301463 W AU0301463 W AU 0301463W WO 2004041812 A1 WO2004041812 A1 WO 2004041812A1
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Prior art keywords
optionally substituted
compound
compound according
group
cells
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English (en)
Inventor
Barbara Martha Meurer-Grimes
Jin Yu
Gino Luigi Vairo
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Sarawak Government of State of Malaysia
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Sarawak Government of State of Malaysia
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Priority claimed from US10/291,863 external-priority patent/US6710075B2/en
Application filed by Sarawak Government of State of Malaysia filed Critical Sarawak Government of State of Malaysia
Priority to EP03769069A priority Critical patent/EP1569928A4/fr
Priority to JP2004548931A priority patent/JP4847014B2/ja
Priority to CA002504999A priority patent/CA2504999A1/fr
Priority to AU2003277988A priority patent/AU2003277988A1/en
Publication of WO2004041812A1 publication Critical patent/WO2004041812A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D407/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00
    • C07D407/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings
    • C07D407/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/06—Antiasthmatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/77—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D307/93—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom ortho- or peri-condensed with carbocyclic rings or ring systems condensed with a ring other than six-membered
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D407/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00
    • C07D407/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings
    • C07D407/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the present invention relates generally to compounds having a cyclopentabenzofuran core. More particularly, the present invention relates to cyclopentabenzofuran compounds having a bulky substituent at the 6-oxy position, such as where the cyclopentabenzofuran core is substituted by a dioxanyloxy moiety. The invention also relates to the use of these compounds in therapy and compositions comprising said compounds.
  • Aglaia is a large genus of the family Meliaceae comprising over 100 (mostly woody) species in Indo-Malaysia and the Western Pacific region. Uses include treatment of fever, fractures, parturition and inflammation. Extracts are also used as bactericides, insecticides, in perfumery, as an astringent, tonic, a refrigerant (Dr Duke's Phytochemical and Ethnobotanical Databases) and for the treatment of abdominal tumours (Pannel, et al, 1992, Kew Bull, (16) 273-283).
  • New compounds have now been isolated from Aglaia leptantha, Miq. (Meliaceae) which uniquely possess a dioxanyloxy group at the 6- position of the cyclopenta[b]benzofuran core.
  • Compounds A and B have been shown to exhibit potent cytotoxic and cytostatic effects on cancer cell growth and viability and thus the compounds of the invention and derivatives thereof, may be useful as therapeutic agents in the treatment of cancer and cancerous conditions or other diseases associated with cellular hyperproliferation.
  • the invention relates to compounds of Formula (I) or a salt or prodrug thereof.
  • each R -R , 10 is independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted acyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted cycloalkylalkyl, optionally substituted arylacyl, optionally substituted cycloalkylacyl and a C-l linked saccharide;
  • X is OR 8 orNR 9 R 10 ;
  • R 11 and R 12 are preferably each independently hydrogen or, alternatively, OR 4 and R 11 , and/or OR 5 and R 12 together form a methylenedioxy group;
  • Y is selected from the group consisting of optionally substituted phenyl, optionally substituted benzyl, optionally substituted benzoyl, optionally substituted C 3 -C 8 cycloalkyl, (preferably optionally substituted C 5 -C 6 cycloalkyl) optionally substituted CH 2 -(C 3 -C 8 cycloalkyl) (preferably optionally substituted CH 2 -(C 5 -C 6 cycloalkyl), optionally substituted 5-6 rnembered heterocyclyl, and optionally substituted CH 2 -(5-6 membered heterocyclyl).
  • the invention relates to compounds (including stereoisomers within the dioxanyl group) of formula (i) or a salt or prodrug thereof.
  • each R ! -R 10 is independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted acyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted cycloalkylalkyl, optionally substituted arylacyl, optionally substituted cycloalkylacyl and a C-l linked saccharide; and X is OR 8 orNR 9 R 10 ;
  • R 11 and R 12 are each independently hydrogen or, OR 4 and R 11 , and/or OR 5 and R 12 together form a methylenedioxy group. In one preferred embodiment, R and R are both hydrogen.
  • compounds of the present invention have the Formula (ii):
  • Formula (ii) has 4 chiral centres in the dioxanyl moiety.
  • Two isomers (isomeric in the dioxanyl group) of Formula (ii) have now been isolated - Compounds A and B as described in Example 1.
  • the invention provides a composition
  • a composition comprising a compound of Formula (I), such as Formula (i), or a salt or prodrug thereof, together with a pharmaceutically acceptable carrier, excipient or diluent.
  • the present invention provides a method for the treatment of cancer or a cancerous condition or a disease state or condition associated with cellular hyperproliferation comprising the administration of a treatment effective amount of a compound of Formula (I), such as Formula (i), or a salt, derivative or prodrug thereof, to a subject in need thereof.
  • a treatment effective amount of a compound of Formula (I), such as Formula (i), or a salt, derivative or prodrug thereof to a subject in need thereof.
  • Some particular cancerous conditions which may be treated by the compounds of the invention may include lung, prostate, colon, brain, melanoma, ovarian, renal and breast tumours and leukemia.
  • Disease states or conditions associated with cellular hyperproliferation which may be treated by compounds of the invention may include atherosclerosis, restinosis, rheumatoid arthritis, osteoarthritis, inflammatory arthritis, psoriasis, peridontal disease or virally induced cellular hyperproliferation.
  • Figure 1 Compound A promotes differentiation of THP-1 leukemic cells.
  • THP-1 cells were cultured for 4 days in the presence or absence of 10 nM Compound A as indicated. Where shown cells were also treated with IFN ⁇ (lOOng/ml) (3 days) or with PMA (0.1 ⁇ M) (4 days) in the presence or absence of Compound A. Images are of cells visualised by phase contrast microscopy (magnification x200).
  • Figure 2 Effects of Compound A on cell cycle progression and viability of THP-1 cells.
  • THP-1 cells were cultured for 2 days with the indicated concentration of Compound A or
  • the numbers indicate the % of cells in the various cell cycle phases relative to all cells with >2N DNA content and also the % dead cells (ie. subdiploid ⁇ 2N cells) to the left of the marker (the vertical line) that arose during the culture period.
  • FIG. 3 Effects of Compound A on the proliferation of A549 cells.
  • A549 cells were seeded at -10,000 cells/well and cultured in the presence of the indicted concentrations of Compound A or paclitaxel. Cells were collected and the viable cell number determined by haemocytometer counting of trypan blue stained cells at the various times. The results are the averages ⁇ SEM of triplicate cultures.
  • Figure 4 Effects of Compound A on cell cycle progression and viability of A549 cells.
  • A549 cells were cultured for 6 days with the indicated concentration of Compound A or 1 ⁇ M paclitaxel then collected and fixed in 70% ethanol prior to staining with propidium iodide and DNA content determined by flow cytometry.
  • the numbers indicate the % of cells in the various cell cycle phases relative to all cells with >2N DNA content and also the % dead cells (ie. subdiploid ⁇ 2N cells) to the left of the marker that arose during the culture period.
  • FIG. 5 Compounds A and B induce G2/M phase accumulation of K562 leukemic cells
  • K562 cells were cultured for 3 days with the indicated concentration of Compounds A or B then collected and fixed in 70% ethanol prior to staining with propidium iodide and DNA content determined by flow cytometry.
  • the numbers indicate the % of cells in G0/G1, S and G2/M phases of the cell cycle respectively relative to all cells with >2N DNA content.
  • A549 cells were seeded at ⁇ 10,000 cells/well and cultured in the presence of the indicted concentrations of Compound A or paclitaxel and the viable cell numbers determined by haemocytometer counting of trypan blue stained cells at the various times. On day 5 some of the cells were washed, resuspended in fresh medium lacking the various treatments and cultured for another 4 days prior to counting.
  • Figure 7 Compound A inhibits camptothecin- and paclitaxel-induced cytotoxicity of A549 cells
  • A549 cells in 6 well plates were cultured for 3 days in the presence or absence of 10 nM Compound A together with 0.1 ⁇ M camptothecin, 10 ⁇ M vinblastin, 1 ⁇ M paclitaxel or 1 ⁇ M staurosporine as indicated.
  • the cells were then collected and fixed in 70% ethanol prior to staining with propidium iodide and DNA content determined by flow cytometry.
  • the numbers indicate the % of cells in the various cell cycle phases relative to all cells with >2N DNA content and also the % dead cells (ie. subdiploid ⁇ 2N cells) to the left of the dotted marker that arose during the culture period.
  • A549 cells were seeded at 10,000 cells/well in 6 well plates in the presence or absence of varying concentrations of Compound A (10 -50 nM) or 250 nM doxorubicin for 10 days prior to their processing and staining overnight for senescence-associated ⁇ -galactosidase activity as described previously (Dimri et al., 1995, Proc Natl Acad Sci USA 1995 92(20):9363-7). For Compound A only the 10 nM treatment is shown but there was no detectable SA- ⁇ gal activity at any other concentrations tested.
  • PC phase contrast microscopy.
  • BF bright field microscopy. Magnification x200.
  • Figure 10 Compound A inhibits growth of human tumour cells in a mouse xenograft model
  • Athymic Balb/c nude mice (Rygard and Povisen, 1969, Ada Pathol Microbiol Scand, 11: 758) were inoculated subcutaneously in the dorsal flank with 2x 10 6 PC3 cells.
  • Compound A was administered (3 mg/kg) after eight days when the tumours became palpable by intraperitoneal injection three times a week.
  • Compound A was first solubilized in ethanol then mixed 1:1 with cremaphore and diluted in saline for injection. Control animals were treated in an analogous manner with the same vehicle but lacking Compound A.
  • the data represents mean tumour volume ⁇ SEM (B) Effect of Compound A on mean tumour mass. At the end of the experiment (29 days post inoculation of PC3 cells) the mice were sacrificed, the tumours excised and then weighed. The data represents mean tumour weight ⁇ SEM.
  • the compounds of the present invention carry a sterically bulky group at the 6-oxy-position, in particular, a dioxanyl group.
  • the dioxanyl group of Formula (ii) (depicted below as sub-Formula (a)) has not previously been reported from a natural source.
  • the invention includes within its scope pharmaceutically acceptable salts, derivatives, or prodrugs of compounds of Formula (I), particularly of Formula (i), such as Compounds A and B.
  • salt, or prodrug includes any pharmaceutically acceptable salt, ester, glycoside, solvate, hydrate or any other compound which, upon administration to the recipient subject is capable of providing (directly or indirectly, for example, by chemical or in vivo enzymatic or hydrolytic degradation) a compound of the invention as described herein.
  • Suitable pharmaceutically acceptable salts include salts of pharmaceutically acceptable inorganic acids such as hydrochloric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic, and hydrobromic acids, or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, toluenesulphonic, benzenesulphonic, salicyclic, sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids.
  • Base salts include, but are not limited to those formed with pharmaceutically acceptable cations, such as sodium, potassium, lithium, calcium, magnesium, ammonium
  • salts can be carried out by methods known in the art. It will also be appreciated that non-pharmaceutically acceptable salts also fall within the scope of the invention, since these may be useful as intermediates in the preparation of pharmaceutically acceptable salts.
  • the compounds of the invention may be in crystalline form or as a solvate (e.g., hydrates). Methods of solvation will be known to those skilled in the art.
  • prodrug includes derivatives that are converted in vivo to the compounds of the invention and include for example, ester (eg acetate) and glycoside derivatives of free hydroxy groups, which may undergo in vivo degradation to release a compound of the invention.
  • ester eg acetate
  • suitable prodrugs may include esters of free carboxylic acid groups.
  • the preparation of suitable prodrugs is further described in Design ofProdrugs, H. Bundgaard, Elseveir, 1985, the contents of which is incorporated by reference.
  • Y groups of Formula (I) in particular the dioxanyl groups of compounds as depicted in Formula (i) and (ii), may possess asymmetric centres and are therefore capable of existing in more than one stereoisomeric form.
  • the invention thus also relates to compounds in substantially pure isomeric form at one or more asymmetric (chiral) centres eg., greater than about 90% ee, such as about 95% or 91% ee, preferably greater than 99% ee, as well as mixtures, including racemic mixtures, thereof.
  • Such isomers may be resolved by conventional methods, eg, chromatography, or use of a resolving agent.
  • the present invention thus provides Compounds A and B.
  • alkyl when used alone or in compound words such as “arylalkyl” refers to a straight chain, branched or cyclic hydrocarbon group, preferably C ⁇ - 2 o, such as C ⁇ - ⁇ o.
  • C ⁇ -C 6 alkyl refers to a straight chain, branched or cyclic alkyl group of 1 to 6 carbon atoms.
  • C ⁇ - 6 alkyl examples include methyl, ethyl, iso- propyl, /.-propyl, «-butyl, -fee-butyl, t-butyl, «-pentyl, isopentyl, 2,2-dimethypropyl, n- hexyl, 2-methylpentyl, 2,2-dimethylbutyl, 3-methylpentyl and 2,3-dimethylbutyl.
  • Examples of cyclic C ⁇ - 6 alkyl include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.
  • alkyl examples include: heptyl, 5-methylhexyl, 1-methylhexyl, 2,2- dimethylpentyl, 3,3-dimethylpentyl, 4,4-dimethylpentyl, 1 ,2-dimethylpentyl, 1,3- dimethylpentyl, 1,4-dimethyl-pentyl, 1,2,3-trimethylbutyl, 1,1,2-trimethylbutyl, 1,1,3- trimethylbutyl, octyl, 6-methylheptyl, 1 -methylheptyl, 1,1,3,3-tetramethylbutyl, nonyl, 1-, 2-, 3-, 4-, 5-, 6- or 7-methyl-octyl, 1-, 2-, 3-, 4- or 5-ethylheptyl, 1-, 2- or 3-propylhexyl, decyl, 1-, 2-, 3-, 4-, 5-, 6-, 7- and 8-methylnonyl, 1-, 2-, 3-, 4-, 5- or 6-e
  • an alkyl group may be optionally substituted by one or more optional substituents as herein defined.
  • the straight, branched or cyclic hydrocarbon group (having at least 2 carbon atoms) may contain one, two or more degrees of unsaturation so as to form an alkenyl or alkynyl group, preferably a C 2 . 2 o alkenyl, more preferably a C 2 - 6 alkenyl, or a C 2 . 2 o alkynyl, more preferably a C . 6 alkynyl. Examples thereof include a hydrocarbon residue containing one or two or more double bonds, or one or two or more triple bonds.
  • alkyl is taken to include alkenyl and alkynyl.
  • aryl when used alone or in compound words such as “arylalkyl”, denotes single, polynuclear, conjugated or fused residues of aromatic hydrocarbons or aromatic heterocyclic (heteroaryl) ring systems, wherein one or more carbon atoms of a cyclic hydrocarbon residue is substituted with a heteroatom to provide an aromatic residue. Where two or more carbon atoms are replaced, this may be by two or more of the same heteroatom or by different heteroatoms. Suitable heteroatoms include O, N, S and Se.
  • aryl examples include phenyl, biphenyl, terphenyl, quaterphenyl, naphthyl, tetrahydronaphthyl, anthracenyl, dihydroanthracenyl, benzanthracenyl, dibenzanthracenyl, phenanthrenyl, fluorenyl, pyrenyl, idenyl, azulenyl, chrysenyl, pyridyl, 4-phenylpyridyl, 3-phenylpyridyl, thienyl, furyl, pyrrolyl, indolyl, pyridazinyl, pyrazolyl, pyrazinyl, thiazolyl, pyrimidinyl, quinolinyl, isoquinolinyl, benzofuranyl, benzothienyl, purinyl, quinazolinyl, phenazinyl,
  • Preferred hydrocarbon aryl groups include phenyl and naphthyl.
  • Preferred heterocyclic aryl groups include pyridyl, thienyl, furyl, pyrrolyl.
  • An aryl group may be optionally substituted by one or more optional substitutents as herein defined.
  • acyl refers to a group -C(O)-R wherein R is any carbon containing moiety such as an optionally alkyl or substituted aryl group.
  • examples of acyl include straight chain or branched alkanoyl such as, acetyl, propanoyl, butanoyl, 2- methylpropanoyl, pentanoyl, 2,2-dimethylpropanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, heptadecanoyl, octadecanoyl, nonadecanoyl and icosanoyl; cycloalkylcarbonyl, such as cyclopropylcarbonyl,
  • acyl is taken to refer to optionally substituted acyl.
  • Optional substituents for alkyl, aryl or acyl include halo (bromo, fluoro, chloro, iodo), hydroxy, C ⁇ - 6 alkyl (eg methyl, ethyl, propyl (n- and i- isomers)), C ⁇ - 6 alkoxy (eg methoxy, ethoxy, propoxy (n- and /- isomers), butoxy (n-, sec- and t-isomers), nitro, amino, C ⁇ - 6 alkylamino (eg methyl amino, ethyl amino, propyl (n- and i- isomers)amino), C ⁇ - 6 dialkylamino (eg dimethylamino, diethylamino, diisopropylamino), halomethyl (eg trifluoromethyl, tribromomethyl, trichloromethyl), halomethoxy (eg trifluoromethoxy, tribromomethoxy, t
  • substituents for Y include, as well as the substituents above, alkyl substituted with one or more of hydroxy C ⁇ - 6 alkyloxy, C ⁇ _ 6 acyloxy, aryloxy, arylC ⁇ - 6 alkyloxy, C ⁇ - 6 cycloalkylC ⁇ - 6 alkyloxy, arylC ⁇ _ 6 acyloxy, C ⁇ - 6 cycloalkylC ⁇ - 6 acyloxy and C 1 -linked saccharidoxy .
  • arylalkyl and cycloalkylalkyl refer to an alkyl group (preferably straight chain) substituted (preferably terminally) by an aryl and a cycloalkyl group, respectively.
  • arylacyl and cycloalkylacyl refer to an acyl group (preferably where R is straight chain alkyl) substituted (for example, terminally substituted) by an aryl and a cycloalkyl group, respectively.
  • Preferred C-l linked saccharides are a furanose or pyranose saccharide (sugar) substituent which is linked to the backbone structure shown in Formula (I) through the saccharides's 1 -carbon (conventional chemical numbering) to form an acetal at any one of positions Ri, R 2 , R , R t , R 5 , R 6 , or R 7 or an ester linkage at the R 8 or an amide at R or Rio position.
  • Exemplary saccharide groups include reducing sugars such as glucose, ribose, arabinose, xylose, mannose and galactoses, each being linked to an oxygen atom of the structure of Formula (I) through the C-l carbon of the saccharide group.
  • a 5-6 membered heterocyclyl group includes aromatic 5-6-membered heterocyclic aryl groups (heteroaryl) as described above and non aromatic 5-6-membered heterocyclic groups containing one or more heteroatoms (preferably 1 or 2) independently selected from O, N, S and Se.
  • heteroatoms preferably 1 or 2 independently selected from O, N, S and Se.
  • examples thereof include dioxanyl, pyranyl, tetrahydrofuranyl, piperidyl, morpholino, piperazinyl, thiomorpholino and saccharides, for example, those described above.
  • each of R 4 -R 7 and R-t-R 7 respectively may independently be selected from the group consisting of hydrogen, methyl, ethyl, ..-propyl, /-propyl, rz-butyl, -fee-butyl, t-butyl, cyclopropylmethyl (or cyclopropylethyl), cyclobutylmethyl (or -ethyl), cyclopentylmethyl (or -ethyl), cyclohexylmethyl (or -ethyl), phenyl, benzyl, acetyl and C-l linked saccharide.
  • the derivatisation of hydroxy groups of Compounds A and B to form compounds of Formula (i), (ie where any one of R'-R 7 is not hydrogen) can be carried out by methods known in the art for alkylating, arylating or acylating hydroxy groups, for example as described in Protective Groups in Organic Synthesis T.W. Greene and P.G.M. Wutz,
  • hydroxy groups can be alkylated using alkyl halides such as methyl iodide or dialkyl sulfates such as dimethyl and diethyl sulfate.
  • Acylation can be effected by treatment with appropriate carboxylic acids, acid halides or acid anhydrides in the presence of a base or coupling agent.
  • Benzylation may be effected by treatment with a benzyl halide compound such as benzyl bromide, chloride or iodide.
  • De-esterification of the methyl ester can be effected by treatment of the ester with aqueous base. Esterification of a carboxylic acid can be achieved by conventional means including treatment with an appropriate alcohol in the presence of acid, or treatment with alkyl sulfates or alkyl halides.
  • Glycosidic formation may be effected chemically, eg by reacting the starting compound with a protected sugar compound in which C-l has been activated by halogenation for coupling with the hydroxyl or carboxyl groups and the sugar hydroxyls have been blocked by protecting groups.
  • glycoside formation may be effected enzymatically using an appropriate glycosyltransferase such as UDP-galactose dependent galactocyltransferase and UDP-glucose dependent glycocyltransferase (SIGMA).
  • Methods for the conversion of a carboxylic acid or ester group; ie. where X is OR 8 to an amide (X is NR 9 R 10 ) are known to the skilled person and may include treatment of a carboxylic acid with an appropriate amine in the presence of a coupling reagent such as DCC or treatment of an acid halide with the appropriate amine.
  • a coupling reagent such as DCC
  • Other methods which may be suitable are described in Larock, R.E, Comprehensive Organic Transformations pp 963- 995, VCH Publishers (1989).
  • protecting group refers to an introduced functionality which may temporarily render a particular functional group, eg hydroxy or carboxylic acid, inactive under certain conditions in which the group might otherwise be reactive.
  • Suitable protecting groups are known to those skilled in the art, for example as described in Protective Groups in Organic Synthesis (supra).
  • Suitable protecting groups for hydroxy include alkyl, (such as C ⁇ -C 6 alkyl), acyl (such as C(O)C ⁇ -C 6 alkyl, benzoyl and the like), benzyl, and silyl groups (such as trimethylsilyl, t-butyldimethyl silyl, t-butyldiphenylsilyl and the like).
  • alkyl, acyl or arylalkyl may serve as either a temporary protecting group or as a non-hydrogen R ⁇ R 8 group in Formula (I).
  • the dioxanyl group may be cleaved from the 6-oxy position of the cyclopentabenzofuran core using known methods to afford a dioxane compound.
  • the resulting dioxane compound could be used to substitute other compounds, such as oxy- substituted compounds, including the corresponding 6-oxy position, or other oxy positions, on other cyclopentabenzofuran compounds such as those described in the references herein.
  • cyclopentabenzofuran compounds having a methoxy substituent at the 6-position, such as those described in the references cited herein (incorporated herein by reference) eg Reference Compounds 1-3 (as described in Example 4), can, where appropriate be 6-demethylated, and the resulting 6-hydroxy group reacted with a suitable Y precursor to form an 6-OY group.
  • Methods therefor are known in the art, for example, one method may involve reacting the 6-OH group with a Y-halogen compound where halogen includes Cl, Br and I.
  • access to the cyclopentabenzofuran core, incorportation of the Y group can be achieved via synthetic methods analogous to that described in Trost et al, J. Am. Chem. Soc, 1990, 112, 9022- 9024.
  • 6-OY compounds form a further aspect of the invention.
  • -R 1 and R 2 are both hydrogen. -R 1 and R 2 are hydrogen, and R 3 is methyl.
  • R 3 -R 5 is methyl, ethyl or propyl, preferably methyl
  • R 3 -R 5 are methyl, ethyl or propyl, preferably methyl
  • R 3 -R 5 are methyl, ethyl or propyl, preferably methyl.
  • R and R are both hydrogen.
  • R 8 is selected from hydrogen, methyl, ethyl or propyl, preferably, methyl.
  • -X is NR 9 R 10 where R 9 and R 10 are both hydrogen or methyl; or R 9 and R 10 are different but at least one of R 9 or R 10 is hydrogen and the other is C ⁇ - 6 alkyl, such as methyl, ethyl or propyl.
  • -Y is an optionally substituted 5-6 membered heterocyclyl group or an optionally substituted C 5 -C 6 cycloalkyl group.
  • Particularly preferred forms of Formula (ii) are Compounds A and B.
  • the compounds of the invention may have use in the treatment of cancerous conditions, or other conditions associated with cellular hyperproliferation, in a subject.
  • Subjects which may be treated by the compounds of the invention include mammals, for example, humans, primates, livestock animals (eg. sheep, cows, horses, goats, pigs), companion animals, (eg. dogs, cats, rabbits, guinea pigs), laboratory test animals, (eg, rats, mice, guinea pigs, dogs, rabbits, primates) or captured wild animals. Most preferably, humans are the subjects to be treated.
  • livestock animals eg. sheep, cows, horses, goats, pigs
  • companion animals eg. dogs, cats, rabbits, guinea pigs
  • laboratory test animals eg, rats, mice, guinea pigs, dogs, rabbits, primates
  • humans are the subjects to be treated.
  • treatment is intended to include the prevention, slowing, interruption or halting of the growth of a cancer, tumour or hyperproliferative cell, or a reduction in the number of targeted cells (or size of the growth mass) or the total destruction of said cell, wherein said cells are cancer, tumour or hyperproliferative cells.
  • Cancerous conditions which may be treated by the compounds of the present invention include conditions wherein the cancers or tumours may be simple (monoclonal, ie composed of a single neoplastic cell type), mixed (polyclonal, ie. composed of more than one neoplastic cell type) or compound (ie. composed of more than one neoplastic cell type and derived from more than one germ layer) and may include benign and malignant neoplasia/hyperplasia.
  • cancerous conditions which may be treated by the present invention include leukemia and breast, colon, bladder, pancreatic, endometrial, head and neck, mesothelioma, myeloma, oesophagal/oral, testicular, thyroid, uterine, prostate, renal, lung, ovarian, cervical brain, skin, liver, bone, bowel and stomach cancers, sarcomas, tumours and melanomas.
  • benign hyperplasias include those of vascular (eg hemangioma), prostate, renal, adrenal, hepatic, colon (eg colonic crypt), parathyroid gland and other tissues.
  • the compounds of the invention may have cytostatic as well as cytotoxic properties, they may also have potential use as therapeutic agents in the suppression of the growth of target populations of cells other than cancer or tumour cells, for example disease states or conditions associated with cellular hyperproliferation.
  • diseases may include atherosclerosis and restinosis (neointimal hyperplasia) and hyperproliferation due to or accompanying an inflammatory response, eg arthritis, (including rheumatoid arthritis, osteoarthritis and inflammatory arthritis), psoriasis and periodontal disease, or cellular hyperproliferation due to the viral infection of cells such as human papilloma virus.
  • the compounds of the invention eg Compounds A and B, may be used in therapy in conjunction with other therapeutic compounds, such as anti-cancer compounds, including paclitaxel, camptothecin, vinblastin and doxorubicin.
  • another aspect of the invention relates to a method for the treatment of cancer or a cancerous condition comprising the administration of an effective amount of a compound of Formula (I) and a further therapeutic agent to a subject in need thereof, and the use of said compound in the manufacture of a medicament for use in conjunction with other therapeutic agents.
  • the compounds of the invention and the further therapeutic agent may be administered simultaneously, as a single composition or as discrete compositions, or may be administered separately, ie, one after the other at suitable intervals as determined by the attending physician.
  • the invention also provides a kit comprising a compound of Formula (I) together with a further therapeutic agent.
  • the term "effective amount" of a compound relates to an amount of compound which, when administered according to a desired dosing regimen, provides the desired therapeutic activity. Dosing may occur at intervals of minutes, hours, days, weeks, months or years or continuously over any one of these periods. Suitable dosages lie within the range of about 0.1 ng per kg of body weight to 1 g per kg of body weight per dosage. The dosage is preferably in the range of 1 ⁇ g to 1 g per kg of body weight per dosage, such as is in the range of 1 mg to 1 g per kg of body weight per dosage. In one embodiment, the dosage is in the range of 1 mg to 500 mg per kg of body weight per dosage.
  • the dosage is in the range of 1 mg to 250 mg per kg of body weight per dosage. In yet another embodiment, the dosage is in the range of 1 ⁇ g to 100 mg per kg of body weight per dosage, such as up to 50 mg per kg body weight per dosage.
  • the dosing regime for each subject may be dependent upon the age, weight, health and medical history of the subject and the extent and progress of the condition to be treated, and can be determined by the attending physician.
  • the active ingredient may be administered in a single dose or a series of doses. While it is possible for the active ingredient to be administered alone, it is preferable to present it as a composition, preferably as a pharmaceutical composition.
  • compositions include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parental (including subcutaneous, intramuscular, intravenous and intradermal) administration.
  • the compositions may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.
  • compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, sachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion.
  • the active ingredient may also be presented as a bolus, electuary or paste.
  • a tablet may be made by compression or moulding, optionally with one or more accessory ingredients.
  • Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g inert diluent, preservative disintegrant such as sodium starch glycolate, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent.
  • a binder e.g inert diluent, preservative disintegrant such as sodium starch glycolate, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose
  • Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.
  • the tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with an enteric coating, to provide release in parts of the gut other than the stomach.
  • compositions suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavoured base, usually sucrose and acacia or tragacanth gum; pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia gum; and mouthwashes comprising the active ingredient in a suitable liquid carrier.
  • compositions for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter, gelatin, polyethylene glycol.
  • compositions suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.
  • compositions suitable for parenteral administration include aqueous and non- aqueous isotonic sterile injection solutions which may contain anti-oxidants, buffers, bactericides and solutes which render the composition isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.
  • the compositions may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use.
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.
  • Preferred unit dosage compositions are those containing a daily dose or unit, daily sub-dose, as herein above described, or an appropriate fraction thereof, of the active ingredient.
  • compositions of this invention may include other agents conventional in the art having regard to the type of composition in question, for example, those suitable for oral administration may include such further agents as binders, sweeteners, thickeners, flavouring agents disintegrating agents, coating agents, preservatives, lubricants and/or time delay agents.
  • suitable sweeteners include sucrose, lactose, glucose, aspartame or saccharine.
  • Suitable disintegrating agents include corn starch, methylcellulose, polyvinylpyrrolidone, xanthan gum, bentonite, alginic acid or agar.
  • 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 may include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc.
  • Suitable time delay agents may include glyceryl monostearate or glyceryl distearate.
  • One or more embodiments of the present invention may also provide methods, compositions agents or compounds which have an advantage over (or avoid a disadvantage) associated with known methods, compositions, agents or compounds used in the chemotherapeutic treatment of cancerous conditions or conditions associated with the hyperproliferation of cells.
  • Such advantages may include one or more of: increased therapeutic activity, reduced side effects, reduced cytotoxicity to non-cancerous or non- proliferative cells, improved solubility or dispersibilty for formulation into pharmaceutical compositions, improved stability or a more readily available means of obtaining said compounds, eg. by simpler, cheaper or higher yielding synthetic or isolation processes.
  • step (f) Chromatograph the concentrate obtained under step (e) on a C-l 8 preparative column (WATERS, Nova-Pak C-l 8, 6 micron, 2.5 x 25 cm) at a flow rate of 20 mL/min using a linear gradient from 25%) to 45% acetonitrile in water in 30 minutes with 0.1%> formic acid.
  • a C-l 8 preparative column WATERS, Nova-Pak C-l 8, 6 micron, 2.5 x 25 cm
  • step (g) Collect and concentrate the eluates with the chromatographic and spectroscopic characteristics outlined in step (d) at approximately 22 minutes.
  • step (h) Chromatograph each eluate obtained under (g) on a Sephadex LH20 column using methanol as a solvent. Collect and concentrate the fractions with spectral characteristics outlined in (d). These samples were used for the structural elucidation of Compounds A and B.
  • the methanol extract obtained under (a) may be partitioned with equal volumes of water and dichloromethane. The dichloromethane phase is then processed according to steps (e) to (h).
  • UV/Vis absorption maxima 223, 275 nm (in MeCN/H 2 O, 0.1% HCOOH).
  • MS Mass spectra were obtained on a Finnigan LCQ iontrap mass spectrometer using the ESI source in the positive ion mode. The sample was dissolved in 0.1 %FA in MeOH and introduced into the source by infusion with a syringe pump at rate of 3 ⁇ L/min.
  • the objective of this experiment was to unambiguously determine the attachment position of the dioxanyl sidechain to the cyclopentabenzofuran core in Compounds A and B.
  • Compounds A and B are cytostatic and cytotoxic for human tumour cell lines
  • TNF- ⁇ Tumour Necrosis Factor- ⁇
  • THP-1 human promonocytic leukemia cells THP-1 human promonocytic leukemia cells activated with lipopolysaccharide (LPS).
  • LPS lipopolysaccharide
  • Compounds A and B potently inhibited TNF- ⁇ production at broadly similar concentrations that were active in the WST-1 reduction, DNA and protein synthesis assays.
  • the effects of Compounds A and B on A549 lung epithelial carcinoma cells were also measured and the data is also included in Table 4.
  • Compounds A and B are significantly less potent for inhibition of interleukin-1 (IL-l)-induced Intercellular Adhesion Molecule-1 (ICAM-1) expression by A549 cells even though in these cells the protein and DNA synthesis inhibition occur at broadly similar concentrations as for THP-1 cells.
  • IL-l interleukin-1
  • IAM-1 Intercellular Adhesion Molecule-1
  • TNF ⁇ TNF ⁇ by THP-1 cells was measured as that released into the culture supernatant over 18 hours by sandwich enzyme-linked immunosorbent assay (ELISA) using the following mouse anti-TNF ⁇ monoclonal antibodies (capture antibody, MAB610; detection antibody, biotinylated MAB210; both from R & D Systems, Minneapolis MN, USA).
  • ICAM-1 Surface expression of ICAM-1 by A549 cells was assayed after 24 hours of culture by direct antibody binding using a europium-labelled mouse anti- ICAM-1 monoclonal antibody (R&D Systems Cat No. BBA3) and measured by time- resolved fluorescence using Delfia assay (EG&G Wallac, Turku, Finland). Reduction of WST-1 (Roche, Cat. No. 1644807) by THP-1 cells was measured after 18 hours of culture according to the manufacturer's instructions.
  • Protein synthesis was measured as the uptake of [ 14 C]-leucine (0.5 ⁇ Ci/mL) after 48 hours for THP-1 cells and 72 hours for A549 cells cultured in growth medium (RPMI-1640, 10%) FBS) containing 10%) the usual L-leucine concentration (5 mg/mL).
  • DNA synthesis was measured as the uptake of [ 14 C]-thymidine (0.5 ⁇ Ci/mL) after 48 hours for THP-1 cells and 72 hours for A549 cells in normal growth medium.
  • Compound A was assessed for cytotoxic and cytostatic activity against a panel of cell lines derived from a variety of human tumour types in addition to THP-1 and A549 cells (Table 5). These included K562 leukemic cells (Lozzio and Lozzio, 1975, Blood 45:321-34), PC3 prostate tumour cells (Kaighn et al, 1979, Invest. Urol. 17:16-23) and SF268 glioblastoma cells (Westphal et al, 1985, Biochem. Biophys. Res. Commun., 132:284-9). Compound A exhibited potent cytostatic activity in nearly all cell lines tested with GI 5 o values ranging between 1-7 nM.
  • Compound A also exhibited potent cytotoxic effects against the various tumour cell lines. Interestingly, the THP-1 and PC3 cells proved the most rapidly killed with little difference in LC 5 o values obtained after 3 or 6 days of culture. However, the cytotoxic potency of Compound A increased dramatically after 6 days of culture for the K562, A549 and SF268 cells. It should be noted that the concentration of Compound A required to inhibit cell proliferation were significantly lower than those required to elicit a cytotoxic response. Hence, the cytostatic effect of Compound A is biochemically distinguishable from its ability to induce cell death. Table 6 shows that Compound B exhibited cytotoxic effects against the various tumour cell lines with comparable potency to that observed with Compound A. Table 5: Compound A has potent cytostatic and cytotoxic activity in various human tumour cell lines in vitro*
  • the GI 50 value represents the concentration of compound that inhibited the cell number increase (relative to untreated cells) by 50% after 3 days of culture. Relative cell number was determined by measuring cellular DNA using a fluorescent DNA-binding dye (YOYO-1) after lysing the cells with digitonin (Becker et al, Anal Biochem, ⁇ 994, 221(l):78-84).
  • the LC 50 value represents the concentration of compound that killed 50%> of the cells.
  • Cell death was measured as the proportion of dead cells exhibiting sub-diploid DNA content determined by flow cytometry after staining with propidium iodide (Nicoletti et al, J. Immunol. Methods, 1991, 139:271-79).
  • Compound A was tested for activity in the National Cancer Institute in vitro anticancer drug discovery screen. For this Compound A was tested at five 10-fold dilutions ranging from 10 "4 M to 10 "8 M against a panel of different human tumour cell lines representing major types of cancer as described by Boyd and Paull, Drug Development Research, 1995, 34:91- 109. Briefly, this involved a 48 hr incubation of the cells with Compound A prior to measuring the relative cell number by staining with sulforhodamine B. GI 50 values represent the concentration of Compound A that inhibited net growth of the cells by 50% compared to untreated controls. LC 50 values represent the concentration of Compound A that resulted in a net 50%) loss (killing) of the cells relative to the start of the experiment. The data represent the average values from two such experiments conducted.
  • Cytotoxic activity of Compound A is not shared by other known related compounds lacking dioxanyloxy substitution.
  • Compound A ' displays cytotoxic activity.
  • Table 8 compares the cytostatic and cytotoxic effects of Compound A to three previously identified lH-cyclopenta[b]benzofuran lignans that lack the dioxanyloxy group at the C6-position.
  • the reference compounds are: Rocaglaol (Reference Compound 1) (Ohse et al, JNat Prod, 1996, 59(7):650-52); 4'-Demethoxy-3',4'-methylenedioxyrocaglaol (Reference Compound 2) and Methyl 4'-demethoxy-3',4'-methylenedioxyrocaglate (Reference Compound 3) (Lee et al, Chem Biol Interact, 1998, 115(3):215-28). All four compounds exhibited detectable cytostatic activity in A549 cells with Compound A being the most potent followed in decreasing order by Reference Compounds 3, 2 and 1 respectively. Importantly, of the compounds tested, other than Compound A none of the Reference
  • GI 50 values were determined by measuring relative changes in cell number using YOYO-1 as described for Table 5.
  • LC 5 o values were determined by measuring cell death as a function of loss of membrane integrity using YOYO-1 uptake (Becker et al, Anal Biochem, ⁇ 994, 221(l):78-84). The structures of the reference compounds are also shown.
  • Table 9 shows that acetylation of the dioxanyl side chain of Compounds A and B did not reduce their biological activity since Compounds A' and B' inhibited WST-1 reduction of THP-1 leukemic with at least similar potencies to the unmodified compounds.
  • the lower IC 5 o values for all the compounds depicted in this WST-1 reduction experiment compared to the values shown in Table 4 reflects the enhanced sensitivity of the cells when treated for 3 days compared to the 18 hr treatment used in the latter assay.
  • Compound A has acute protein synthesis inhibitory activity
  • THP-1 cells and A549 cells were pretreated with the indicated concentrations of Compound A for 1 hour prior to the addition of (1 ⁇ Ci/mL) [ 14 C] leucine (protein synthesis) or [ 14 C] thymidine (DNA synthesis) for a further 2 hours.
  • the IC 5 o values represent the concentration of Compound A required to inhibit incorporation of isotope by 50%) relative to untreated control cell cultures.
  • Compound A induces differentiation of human leukemic cell lines.
  • This widely used cell line is well characterised as a model of human myelomonocytic differentiation (Collins, Blood, 1987, 70(5): 1233-44).
  • monocytic differentiation was quantitated by measuring CD 14 surface antigen expression by flow cytometric analysis.
  • CD14 an LPS-binding protein, is expressed on the surface of cells of the myelomonocytic lineage and is normally expressed at very low levels in undifferentiated HL60 cells (Ferrero et al., R/ooi,1983,61(l):171-9).
  • Table 10 shows that Compound A at doses greater than 10 nM significantly enlianced CD 14 expression in the viable HL60 cells remaining after 4 days of culture. Taken together these data strongly indicate that Compound A has the ability to induce differentiation of human leukemic cell lines.
  • HL60 cells were cultured for 4 days with the indicated concentration of Compound A then collected and fixed in 70% ethanol.
  • Cells were then stained with mouse monoclonal anti CD14 antibody (OKM1) and this was measured using FITC-conjugated goat anti-mouse IgGl as a secondary antibody. Stained cells were visualised by flow cytometry and analysis was restricted to cells judged viable at the time of fixing based on their forward and side light-scatter characteristics. Non specific staining of cells was controlled for by incubating with secondary antibody only.
  • Cytostatic activity of Compound A is associated with a general inhibition of cell cycle progression in A549 cells
  • FIG. 2 shows that the microtubule destabilising drug paclitaxel (Sorger et al, Curr Opin Cell Biol, 1997, 9(6):807-14) which also induced THP-1 cell death, caused cells to accumulate in the G2/M phases of the cell cycle.
  • Compound A inhibits cell cycle-dependent cytotoxicity elicited by various anti-cancer agents
  • Camptothecin is an inhibitor of DNA topoisomerase 1, an enzyme required for DNA replication, and results in pertubation of the S phase of the cell cycle with subsequent cell death due to activation of an S phase checkpoint (Darzynkiewicz et al, Ann N Y Acad Sci, 1996, 803:93-100).
  • Paclitaxel inhibits microtubule function required for formation of the mitotic spindle thereby resulting in activation of an M phase checkpoint and subsequent cell death (Sorger et al, Curr Opin Cell Biol, 1997 9(6): 807- 14).
  • Figure 7 shows that 10 nM Compound A significantly reduced the cytotoxic effects of both camptothecin and paclitaxel even when these drugs were added at up to a 2000-fold excess. Compound A may, in a dominant manner, prevent the cell cycle-dependent cytotoxic effects of camptothecin and paclitaxel.
  • camptothecin resulted in accumulation of cells in S phase of the cell cycle and also increased the level of dead cells detected as those with a sub-diploid DNA content.
  • both vinblastin and paclitaxel resulted in the majority of cells arresting in the G2/M phases of the cell cycle and increased appearance of sub-diploid dead cells.
  • the presence of 10 nM Compound A prevented their characteristic cell cycle arrest and significantly inhibited their cytotoxic effects, dramatically reducing the appearance of sub-diploid dead cells.
  • Compound A had little effect on the cytotoxic effects of staurosporine, an agent which appears capable of killing cells at all active phases of the cell cycle.
  • Cytostatic effects of Compound A do not correlate with a biomarkerfor replicative senescence.
  • SA- ⁇ -gal activity a biomarker previously described to correlate well with senescence of human cells.
  • SA- ⁇ -gal activity a biomarker previously described to correlate well with senescence of human cells.
  • some anti-cancer agents that act by diverse mechanisms including doxorubicin, cisplatin, cytarabine, etoposide and paclitaxel, can all induce SA- ⁇ -gal activity in a variety of tumour cell lines (Chang et al, Cancer Res 1999, 59(15):3761-7).
  • Compound A inhibits cell proliferation but not increased cell size
  • Compound A inhibits growth of human tumour cell lines in a mouse xenograft tumour model.

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Abstract

La présente invention concerne des composés comprenant un noyau de cyclopentabenzofurane, et l'utilisation de ces composés en thérapie, ainsi que des compositions contenant lesdits composés.
PCT/AU2003/001463 2002-11-08 2003-11-06 Composes et methodes therapeutiques Ceased WO2004041812A1 (fr)

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CN115385924A (zh) * 2022-10-06 2022-11-25 贵州省中国科学院天然产物化学重点实验室(贵州医科大学天然产物化学重点实验室) 一种具有抗肿瘤活性的环戊烷苯并呋喃类化合物及其应用
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EP3597184A3 (fr) * 2011-06-16 2020-04-22 Korea Research Institute of Chemical Technology Dérivés d'indanone, sels pharmaceutiquement acceptables ou isomères optiques correspondants, procédé de préparation de ceux-ci et compositions pharmaceutiques les contenant en tant qu'ingrédient actif pour la prévention ou le traitement de maladies virales
US9464067B2 (en) 2011-06-16 2016-10-11 Katholieke Universiteit Leuven K.U. Leuven R & D Indanone derivatives, pharmaceutically acceptable salts or optical isomers thereof, preparation method for same, and pharmaceutical compositions containing same as active ingredient for preventing or treating viral diseases
EP2722042A4 (fr) * 2011-06-16 2015-04-22 Korea Res Inst Chem Tech Dérivés indanone, leurs isomères optiques ou leurs sels pharmaceutiquement acceptables, leur procédé de préparation, et des compositions pharmaceutiques les contenant en tant que principe actif pour prévenir ou traiter des maladies virales
US9890133B2 (en) 2011-06-16 2018-02-13 Katholieke Universiteit Leuven K.U. Leuven R&D Indanone derivatives, pharmaceutically acceptable salts or optical isomers thereof, preparation method for same, and pharmaceutical compositions containing same as active ingredient for preventing or treating viral diseases
US9790197B2 (en) 2012-12-14 2017-10-17 Katholieke Universiteit Leuven K.U. Leuven R & D Compound, pharmaceutically acceptable salt or optical isomer thereof, method for preparing the same, and pharmaceutical composition for prevention or treatment of viral diseases containing same as active ingredient
US9951058B2 (en) 2012-12-14 2018-04-24 Katholieke Universiteit Leuven K.U. Leuven R & D Compound, pharmaceutically acceptable salt or optical isomer thereof, method for preparing the same, and pharmaceutical composition for prevention or treatment of viral diseases containing same as active ingredient
WO2017214024A1 (fr) 2016-06-06 2017-12-14 Genentech, Inc. Médicaments conjugués d'anticorps silvestrol et procédés d'utilisation
US10639378B2 (en) 2016-06-06 2020-05-05 Genentech, Inc. Silvestrol antibody-drug conjugates and methods of use
US11648315B2 (en) 2016-06-06 2023-05-16 Genentech, Inc. Silvestrol antibody-drug conjugates and methods of use
US12083092B2 (en) 2020-04-20 2024-09-10 Novartis Ag Antiviral 1,3-di-oxo-indene compounds
US12286423B2 (en) 2020-04-20 2025-04-29 Novartis Ag Antiviral 1,3-di-oxo-indene compounds
CN115385924A (zh) * 2022-10-06 2022-11-25 贵州省中国科学院天然产物化学重点实验室(贵州医科大学天然产物化学重点实验室) 一种具有抗肿瘤活性的环戊烷苯并呋喃类化合物及其应用
CN115385924B (zh) * 2022-10-06 2023-10-13 贵州省中国科学院天然产物化学重点实验室(贵州医科大学天然产物化学重点实验室) 一种具有抗肿瘤活性的环戊烷苯并呋喃类化合物及其应用

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AU2003277988A1 (en) 2004-06-07
CA2504999A1 (fr) 2004-05-21

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