EP1150967A1 - Therapeutic compounds and methods - Google Patents
Therapeutic compounds and methodsInfo
- Publication number
- EP1150967A1 EP1150967A1 EP99970658A EP99970658A EP1150967A1 EP 1150967 A1 EP1150967 A1 EP 1150967A1 EP 99970658 A EP99970658 A EP 99970658A EP 99970658 A EP99970658 A EP 99970658A EP 1150967 A1 EP1150967 A1 EP 1150967A1
- Authority
- EP
- European Patent Office
- Prior art keywords
- group
- compound
- alkyl
- alkylamino
- alkyl group
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Withdrawn
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C69/00—Esters of carboxylic acids; Esters of carbonic or haloformic acids
- C07C69/66—Esters of carboxylic acids having esterified carboxylic groups bound to acyclic carbon atoms and having any of the groups OH, O—metal, —CHO, keto, ether, acyloxy, groups, groups, or in the acid moiety
- C07C69/73—Esters of carboxylic acids having esterified carboxylic groups bound to acyclic carbon atoms and having any of the groups OH, O—metal, —CHO, keto, ether, acyloxy, groups, groups, or in the acid moiety of unsaturated acids
- C07C69/734—Ethers
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C59/00—Compounds having carboxyl groups bound to acyclic carbon atoms and containing any of the groups OH, O—metal, —CHO, keto, ether, groups, groups, or groups
- C07C59/40—Unsaturated compounds
- C07C59/58—Unsaturated compounds containing ether groups, groups, groups, or groups
- C07C59/64—Unsaturated compounds containing ether groups, groups, groups, or groups containing six-membered aromatic rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D317/00—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms
- C07D317/08—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3
- C07D317/44—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D317/46—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems condensed with one six-membered ring
- C07D317/48—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring
- C07D317/50—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to atoms of the carbocyclic ring
- C07D317/60—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
Definitions
- the present invention generally contemplates compounds which are useful in therapy. More particularly, the present invention relates to compounds which are useful in the treatment of cancer. In particular, the present invention relates to bi-aryl compounds and their use in the treatment of cancer.
- Taxol ® and analogues which contain the fused tricyclic taxane ring system. Since its first isolation in the 1960's from the bark of the Pacific Yew tree, Taxol ® and its analogues have been viewed as leading target candidates in the search for an effective treatment for cancer, in particular breast and ovarian cancer. However, whilst demonstrating significant cytotoxic activity, Taxol ® itself, and the taxanes in general, are poorly soluble in the solvents considered acceptable for human administration. This has prompted sigmficant activity in the search for more soluble second generation derivatives, and for suitable formulation processes which provide pharmaceutical preparations which can effectively administer the required therapeutic dosages. Furthermore, although synthetic routes to the taxanes have been developed, their challenging structural complexity has thus far dictated that natural rather than synthetic sources remain the most viable means of obtaining commercially effective quantities for medical use, albeit by semi-synthetic routes.
- the present invention provides for use in therapy a compound according to one of Formulae:
- each R is independently selected from, a C, .10 alkyl group, a phenyl
- C,_ 10 alkyl group a hydroxy group, a C 0 alkoxy group, a phenyl C l 0 alkoxy group, an amino group a C,_ 6 alkylamino group, a di-C 0 alkylamino group, a halogen atom, a -O- C(O)-C, .10 alkyl group, a -C(O)-C .10 alkyl group, a formyl group, a carboxy group, a carboxyC j.jQ alkylester, a carboxamido group, or where p and/or q are greater than 1, any two adjacent R groups together may optionally form the group -W-CH 2 -Z- where W and Z are independently selected from O or NH; each A may be the same or different and is a group CO 2 Q wherein Q is selected from hydrogen, C,-C 10 alkyl, phenyl C r C 10 alkyl, amino, C,-C
- B is a group selected from a hydrogen atom, a hydroxy group, a C 0 alkyl group, a C 0 alkoxy group, an amino group a C x _
- the present invention provides a method of treating cancer comprising the administration of an effective amount of a compound according to any one of Formulae I-IV to a subject in need thereof.
- the present invention provides a composition comprising a compound according to any one of Formulae I-IV together with a pharmaceutically acceptable carrier or excipient.
- the present invention provides an agent for the treatment of cancer, said agent comprising a compound according to any one of Formulae I-IV.
- the present invention also provides a novel compound according to any one of general Formulae I-IV or a pharmaceutically acceptable derivative thereof.
- the compound is selected from Formula I or II, preferably from Formula I.
- Figure 1 graphically depicts the effect of Compound I-A on human cell lines.
- Figures 2-5 graphically depict dose response curves of Compound I-A, Taxol and Colchicine with Hep G2/A2 and Hep 3B/T2 cell lines in serum-free and culture media.
- Figure 6 graphically depicts the cytotoxicity of Compound I-A, Taxol and Colchine on human fibroblast cells.
- FIGS 7-10 graphically depict cell survival dose response curves following removal of the administered drug ( Compound I-A, Taxol and Colchine) after 2 days.
- Figures 11-12 graphically depict cell cycle analysis of Hep G2/A2 and Hep 3B/T2 cell lines when treated with Compound I-A, Taxol or Colchine.
- Figures 13-14 photographically depict the DNA fragmentation analysis of Hep G2/A2 and Hep 3B/T2 eel lines treated with Compound I-A and Taxol.
- FIGS 15-16 illustrate the micronucleation in Hep G2/A2 and Hep 3B/T2 cells treated with Compound I-A and Taxol.
- Figure 17 depicts an immunoblot of Hep G2/A2 and Hep 3B/T2 cells treated with Compound I-A, Taxol and Colchicine.
- Figures 18-21 graphically depict the weight and volume of mmours in nude mice following implantation of Hep 3B/T2 cells and treatment with Compound I-A or Taxol.
- Figure 22 depicts the effects of Compound I-A and Taxol on microtable polymerization.
- Figure 23 graphically deptics the ability of Compound I-A and Taxol to promote microtubule polymerization as monitored by change in absorbance at 350nm.
- Figure 24 depicts the effect of Compound I-A, taxol or colchicine tubule polymerization.
- the present invention provides for use in therapy, particularly in the treatment of cancer, a compound according to one of formulae:
- each R is independently selected from, a C 0 alkyl group, a phenyl
- each A may be the same or different and is a group CO 2 Q wherein Q is selected from hydrogen, C j -C ⁇ alkyl, phenyl C ⁇ -C 10 alkyl, amino, C j -C 10 alkylamino or di-C,-C 10 alkylamino; or
- the two A groups together form a group -Y-X-C(O)-, wherein Y is selected from C(O) or (CH 2 )n, with n selected from 0, 1 or 2, and X is selected from CH 2 , NH or oxygen;
- B is a group selected from a hydrogen atom, a hydroxy group, a
- the therapeutic use is in the treatment of cancer in mammals, for example, humans, primates, livestock animals (eg. sheep, cows, horses, goats, pigs), companion animals (eg. cats, dogs), laboratory test animals (eg. mice, rats, guinea pigs, rabbits) or captured wild animals. More preferably, the therapeutic use is in the treatment of cancer in humans.
- mammals for example, humans, primates, livestock animals (eg. sheep, cows, horses, goats, pigs), companion animals (eg. cats, dogs), laboratory test animals (eg. mice, rats, guinea pigs, rabbits) or captured wild animals.
- the therapeutic use is in the treatment of cancer in humans.
- the present invention also provides a method of treating cancer comprising the administration of an effective amount of a compound according to any one of Formulae I-IV, or a pharmaceutically acceptable derivative, to a subject in need thereof.
- the cancers or tumours which may be treated by the compounds, compositions and methods of the invention may simple (monoclonal, i.e., composed of a single neoplastic cell type), mixed (polyclonal, i.e., composed of more than one neoplastic cell type) or compound (i.e., composed of more than one neoplastic cell type and derived from more than one germ layer).
- Some examples of cancers which may be treated by the invention include breast, colon, uterus, prostate, lung, ovarian, skin, liver and stomach cancers, tumours and melanomas.
- C ⁇ - 10 alkyl when used in the terms "C, .10 alkyl group”, “C 0 alkylamino group”, “di-C ,. 10 alkylamino group”, phenyl C , .10 , carboxy C 0 alkylester, "-C(O)-C 0 alkyl” and “-O-C(O)-C 0 alkyl” refers to an alkyl (or alkylene) group, which contains 1-10 carbon atoms, preferably 1 to 6 carbon atoms, ie.
- methyl, ethyl, propyl, butyl, pentyl or hexyl and includes, where applicable, branched and cyclic alkyl groups as well as straight chain alkyl groups. Particularly preferred alkyl groups are C,_ 4 alkyl groups, particularly methyl, ethyl and propyl.
- a "phenyl C r C 10 alkyl” group refers to a phenyl group linked by a C,-C 10 alkylene chain.
- C,_ 10 alkoxy is intended to refer to C _ l0 alkyl group as defined above, when linked by an oxygen atom.
- halogen refers to fluorine, chlorine, bromide or iodine.
- phenyl As used herein the terms "phenyl”, “amino”, “carboxamido” and “alkyl” as used alone or in the terms “alkoxy”, “alkylamino”, “dialkylamino”, “phenylalkyl”, “carboxylalkyl ester”, "-C(O)-alkyl” (acyl) and “-O-C(O)-alkyl” (acyloxy) also refer to these groups where they can be optionally substituted by one or more substituents eg. "alkyl” refers to optionally substituted alkyl.
- Suitable optional substituents include, hydroxy, alkyl, halo, phenyl, alkoxy, amino, alkylamino, dialkyl-imino, nitro, acyl, acyloxy, carboxy, carboxy ester, carboxamido, formyl, cyano, nitro, sulphate, phosphate.
- Optional substimtion of "alkyl” is also intended to refer to one or more degrees of unsaturation i.e. one or more double or triple bonds, so as to form
- the two A groups together form a group -Y-X-C(O)-.
- X is NH or oxygen. Even more preferably, X is oxygen.
- Y is (CH 2 )n. More preferably n is 1.
- a particularly preferred embodiment of -Y-X-C(O)- is -CH 2 -O-C(O)- or -C(O)-O-C(O)-.
- A is the group -CO 2 Q.
- Q include: H, CH 3 , CH CH 3 (CH 2 ) 2 CH 3 (CH 2 ) g CH 3 (CH 2 ) 6 CH 3 (CH 2 ) 7 CH 3 (CH 2 ) 8 CH 3 CH(CH 3 ) 2 CH 2 CH(CH 3 ) 2 CH 2 CH 2 CH(CH 3 ) 2 CH 2 Ph, (CH 2 ) 2 Ph, and (CH 2 ) 3 Ph, preferably with Q being an alkyl chain of at least C 2 , or an alkylene-phenyl group of at least C 2 -phenyl.
- the compounds for use in the present invention contain the moiety (i):
- each R group is independently selected from C,_ 4 alkyl, hydroxy, C alkoxy, eg., methoxy, ethoxy, propoxy (n- and iso-), butoxy (/--, sec- and t-), benzyloxy or acetoxy or, where p and/or q is greater than 1, two adjacent R groups may together form the group -O-CH 2 -O-.
- a preferred value for p and q is 1, 2 or 3.
- the -O-CH 2 -O- group may reside on the phenyl group at the 2,3- or 3,4- positions.
- R groups when two adjacent R groups together form the group -O-CH 2 -O-, this is substituted on the phenyl group at the 3,4- positions.
- R groups when p and/or q are 1, 2 or 3, the R groups reside on the benzene rings at the 3-, 4- or 5- positions.
- a preferred subgroup of Formual I has the structure formula F
- I-E Q (CH 2 ) 2 CH 3
- I-F Q (CH 2 ) 3 CH 3
- I-G Q (CH 2 ) 4 CH 3
- I-H Q (CH 2 ) 5 CH 3
- I-I Q (CH 2 ) 6 CH 3
- I-J Q (CH 2 ) 7 CH 3
- I-K Q (CH 2 ) 8 CH 3
- I-L Q CH(CH 3 ) 2
- I-M Q CH 2 CH(CH 3 ) 2
- a particularly preferred compound for use in the present invention is Compound I-A which is isolated from the heartwood extracts of Taiwania cryptomeriodies Hayata.
- Compound I-A may be obtained from the natural source as described in the Examples.
- the compounds of Formula 1-B may be prepared by condensation of piperonal with a succinate such as diethyl succinate in the presence of a base such as sodium methoxide.
- Compounds of Formula I-C to I-Q may then be prepared by appropriate esterification of the carboxylic acid groups.
- amidation of the carboxylic acid groups, or where appropriate the ester group, with the appropriate amine will afford the compounds where A is CO 2 NR'R" wherein R' and R" are independently selected from hydrogen, or C 0 alkyl.
- Methods of amidation are known to the skilled person (see for example Advanced Organic Chemistry Reactions, Mechechanisms, and Structure; 3rd Edition, Jerry March, Wiley Interscience).
- the compound I-R can be formed by treating I-B with a suitable dehydrating agent such as acetyl chloride.
- a suitable dehydrating agent such as acetyl chloride.
- Compound I-S is also known as savinin, or Taiwanin B, and is also isolable from Taiwania cryptomerioides Hayata.
- R groups may be effected by replacing piperonal with an the appropriately substituted benzaldehyde.
- compounds I-T to I-W can be prepared using the appropriate methoxy and benzyloxy substituted benzaldehyde, optionally protected by one or more protecting groups.
- the substiuents on the benzaldehyde may be optionally protected by one or more protecting groups which may be removed at a later stage if desired.
- Suitable protecting groups are known to the skilled person and examples are described in Protective Groups in Organic Synthesis by T. W. Greene & P. Wutz, 2nd Edition (1991), John Wiley and Son.
- the benzaldehyde may be substituted by one or more groups (synthons) which may at a later stage be converted into the desired R group(s) by one or more synthetic transformations.
- groups which may be convertible into suitable R groups are known to the skilled person and are described in standard reference texts e.g., March (supra) and Comprehensive Organic Transformations, Richard C. Larock, V.C.M. Publishers (1989).
- the R group(s) (or the synthon therefor) may be incorporated into the molecule after condensation of a suitable benzaldehyde with the succinate, for example by electrophilic aromatic substimtion or Friedel-Crafts acylation.
- Imides may be formed from the appropriate anhydride by reaction with ammonia (see for example March supra).
- pharmaceutically acceptable derivative refers to any pharmaceutically acceptable salt, ester, solvate, hydrate or any other compound which, upon administration to the recipient is capable of providing (directly or indirectly) a compound 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, maleic, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, toluenesulphonic, benezenesulphonic, salicyclic sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric
- the compounds as defined by Formulae I-IV are administered to the subject in need of therapeutic treatment in an effective amount.
- the term "effective amount” 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 about 10 g per kg of body weight per dosage. The dosage is preferably in the range of 1 ⁇ g to 10 g per kg of body weight per dosage. More preferably, the dosage is in the range of 1 mg to 10 g per kg of body weight per dosage.
- the dosage is in the range of 1 mg to 5 g per kg of body weight per dosage. In another preferred embodiment, the dosage is in the range of 1 mg to 2 g per kg of body weight per dosage. In yet another preferred embodiment, the dosage is in the range of 1 mg to 1 g per kg of body weight per dosage.
- 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.
- the present invention provides a composition comprising a compound according to any one of Formulae I-IV together with a pharmaceutically acceptable carrier or excipient.
- the carrier or excipient must be pharmaceutically "acceptable” in the sense of being compatible with the other ingredients of the composition and not injurious to the subject.
- 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.
- 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.
- 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.
- 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 include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc.
- Suitable time delay agents include glyceryl monostearate or glyceryl distearate.
- An example of a carrier for formulating the compounds for use in the present invention is 1,2-propanediol/ethanol (1 :1 v/v).
- the present invention also provides novel compounds according to any one of general Formulae I-IV as defined above or a pharmaceutically acceptable derivative thereof.
- 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.
- Such advantages may include one or more of increased therapeutic activity, reduced side effects, reduced cytotoxicity to non- cancerous cells, improved solubility or dispersibility for formulation into pharmaceutical compositions, improved stability or a more readily available means of obtaining said compound, eg. by simpler or higher yielding synthetic processes.
- Human cells were seeded in 24-well plates at a density of 10 6 cells/well in DMEM(RPM11640) medium containing 10% fetal calf serum. After 24h incubation, cells were washed three times with phosphate buffered saline (pH 7.0) and treated with various concentration of drags in serum-free DMEM for 2, 4, 6 days. The medium was changed every two days. The viable cells in each well were determined by trypan blue exclusion and haemocytometer counting.
- Cells were treated with serum-free DMEM alone (control), plus drugs for 48 hours and harvested to be permeated by acetone followed by fixing with 3% formaldehyde in phosphate buffered saline (PBS). Nuclei of the fixed cells were incubated with first antibody in 10% BSA phosphate buffered saline for 1 hour in covered glass, washed with PBS buffer three times, followed by a second antibodies incubation and then stained with Hoechst dye 33258 (1 ⁇ g/ml) for immunofluoresent counting and photograph.
- PBS phosphate buffered saline
- the six weeks old male Balb/c-nu/nu nude mice were obtained from national animal center which is charged by the National Science Coucil, Taipei. The obtained nude mice were stabilized in the animal room for two weeks. Hep3B cells which would transform into tumour, at 1 x 10 7 cells/mouse were then injected subcutaneously (S-C) into nude mice. After 5 days of Hep3B cells implanted into nude mice, various dose of drugs or saline (control) were given by I.P., three times/week for continuous three weeks. Those experimental nude mice were kept for another three weeks then the animal was sacrificed and the weight of the tumour measured. The tumour size of the animal were measured every week.
- Dose response curves of Compound I-A with human cell lines (Hep G2/A2, Hep 3B/T2, SCM-1 (stomach), HuH-7 (liver), H1299 (lung) and fibroblast (human normal skin)) are depicted in Figures l(a)-(c).
- Dosages were administered at concentrations of 0-8 ⁇ M and cell numbers were determined against a control (no Compound I-A) after 2 days (b) and 6 days (c).
- Compound I-A The cytotoxicity of Compound I-A was compared against that of the known anti- cancer drugs Taxol and Colchicine. Each compound was administered at concentrations of 0-7.5 ⁇ M and cells were counted at 2,4,6 and 8 days.
- Figures 4 and 5 depict the dose response curves of Compound I-A, Taxol and Colchicine administered in culmre medium (with sera) for Hep G2/A2 and Hep 3B/T2 cell lines respectively.
- the active compounds were administered at concentrations of 0-2.5 ⁇ M and cell survival was monitored at 2,4 and 6 days.
- Figure 6 shows that Compound I-A was, generally less cytotoxic to normal non- cancerous cells than Taxol or Colchicine.
- the dosages of Compound I-A, Taxol and Colchicine were 2.5 ⁇ M, 1.1 ⁇ M and 2.5 ⁇ M respectively.
- Cell death by apoptosis is illustrated by DNA fragmentation, Micronucleation Staining, Nuclear Mitotic Apparatus Protein (NuMA) analysis.
- NuMA Nuclear Mitotic Apparatus Protein
- HepG2/A2 and Hep 3B/T2 cell lines were culmred in serum free and culmre medium.
- Compound I-A (Taiwanin A) was administered at 0.25 ⁇ M and 2.5 ⁇ M and compared against Taxol (1.1 ⁇ M). Analysis was performed 48 hours after the addition of the drugs. The results are illustrated in Figures 13 and 14.
- Compound I-A was administered at 2.5 mg/kg, 500 ⁇ g/kg and 100 ⁇ g/kg and weight and volume of the mmours was recorded at the first administration of the drag and every 5 days for 3 weeks.
- Taxol (10 mg/kg) was monitored for comparision.
- HepG2/A2 cells were left untreated (Ctl) or were treated with 1.5 ⁇ M Taxol or 2.5 ⁇ M Taiwanin-A for 24 hours. Cells were collected and processed for quantitation of tubulin polymerization. The soluble (S) and polymerized (P) fractions were analyzed by immunoblotting with anti- ⁇ -tubulin antibody.
- MAP-depleted tubulin were purified from porcine brain.
- Each reaction mixture (0.15 ml) contained lmg/ml of tubulin, 0.1M MES bufer (0.1 M MES, lmM EGTA, lmM MgCl 2 , pH6.8) and 10 ⁇ M taxol or 2.5 ⁇ M Compound I-A in the presence of GTP at 37 °C.
- Microtubule polymerization was monitored by change of absorbance at 350nm.
- I-B was dehydrated with acetyl chloride to give ⁇ , ⁇ '-dipipero-nylidenesuccinic anhydride (I-R).
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Abstract
Description
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Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US10488598P | 1998-10-20 | 1998-10-20 | |
US104885P | 1998-10-20 | ||
PCT/AU1999/000908 WO2000023432A1 (en) | 1998-10-20 | 1999-10-19 | Therapeutic compounds and methods |
Publications (2)
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EP1150967A1 true EP1150967A1 (en) | 2001-11-07 |
EP1150967A4 EP1150967A4 (en) | 2003-03-05 |
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EP99970658A Withdrawn EP1150967A4 (en) | 1998-10-20 | 1999-10-19 | Therapeutic compounds and methods |
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EP (1) | EP1150967A4 (en) |
AU (1) | AU1140500A (en) |
WO (1) | WO2000023432A1 (en) |
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CN112300104B (en) * | 2020-11-26 | 2022-05-20 | 辽宁中医药大学 | Lignanoid compound in purslane and extraction and separation method and application thereof |
Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
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WO2000010991A1 (en) * | 1998-08-25 | 2000-03-02 | Yale University | Inhibition and treatment of hepatitis b virus and flavivirus by helioxanthin and its analogs |
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WO1988003805A1 (en) * | 1986-11-19 | 1988-06-02 | Chemex Pharmaceuticals, Inc. | Pharmacologically active compounds and mixtures thereof, organic compositions and metal salts |
EP0797564B1 (en) * | 1995-10-18 | 2001-06-27 | Kanoldt Arzneimittel Gmbh | Lignans, a process for their production and pharmaceutical compositions and uses thereof |
-
1999
- 1999-10-19 EP EP99970658A patent/EP1150967A4/en not_active Withdrawn
- 1999-10-19 AU AU11405/00A patent/AU1140500A/en not_active Abandoned
- 1999-10-19 WO PCT/AU1999/000908 patent/WO2000023432A1/en not_active Application Discontinuation
Patent Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2000010991A1 (en) * | 1998-08-25 | 2000-03-02 | Yale University | Inhibition and treatment of hepatitis b virus and flavivirus by helioxanthin and its analogs |
Non-Patent Citations (1)
Title |
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See also references of WO0023432A1 * |
Also Published As
Publication number | Publication date |
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AU1140500A (en) | 2000-05-08 |
EP1150967A4 (en) | 2003-03-05 |
WO2000023432A1 (en) | 2000-04-27 |
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