Classifications

• • 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
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
  • A61K31/7048—Compounds having saccharide radicals and heterocyclic rings having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin, digitoxin or digoxin
• • 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
  • A61K31/365—Lactones
  • A61K31/366—Lactones having six-membered rings, e.g. delta-lactones
  • A61K31/37—Coumarins, e.g. psoralen
• • 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
  • A61P31/20—Antivirals for DNA viruses
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Definitions

• the chaperone protein is Hsp90, which is one of the most abundant proteins in eukaryotic cells, comprising 1-2% of total cellular protein, even under non-stress conditions.
• Hsp90 isomers in the cytosol
• Hsp90 ⁇ and Hsp90 ⁇ in humans
• Hsp86 and Hsp84 in mice
• a third homologue glucose regulated protein 94 (Grp94)
• Rhp94 glucose regulated protein 94
• Hsp75 An additional truncated, cytosolic member of the family, designated Hsp75, has recently been identified.
• Derivatives include, e.g., alkyl, hydroxy, and methoxy derivatives, as well as more complicated derivatives, such as, e.g., 3,4- dihydrocoumarin, 6-methylcoumarin, umbelliferone (7-hydroxycoumarin), 4- hydroxycoumarin (shown below), dicumarol (shown below), warfarin (3 -substituted 4-hydroxycoumarins, an example of which is shown below), phenprocoumon (shown below), coumarone (benzofuran), and coumarone-indene resins.
• R 1 and R 2 can be any suitable substituent.
• R 5 is an aryl, more preferably, monocyclic, and most preferably, 6-membered and substituted with, e.g., halogen (e.g., fluorine, chlorine, or bromine), alkyl, haloalkyl, alkenyl, alkynyl, alkoxy, haloalkoxy, nitro, cyano, hydroxy, amino, thio, alkylthio, or acyl.
• halogen e.g., fluorine, chlorine, or bromine
• substituents may, themselves, be substituted or unsubstituted.
• alkylthio means alkyl, as defined herein, which has a sulfur substituent.
• alkylthios include methanethiol, ethanethiol, and the like.
• alkoxy means a straight-chain or branched-chain alkoxy radical, which has one or more ether groups of the general formula O-R and contains from about 2 to about 20 carbon atoms, preferably from about 2 to about 10 carbon atoms, more preferably from about 2 to about 8 carbon atoms, and most preferably from about 2 to about 6 carbon atoms.
• alkoxy radicals include methoxy, ethoxy, and the like.
• heterocycle and heterocyclic refers to both heterocycloalkylsand heteroaryls.
• heterocycloalkyl means a cycloalkyl radical as defined herein (including polycyclics), wherein at least one carbon of a carbocyclic ring is substituted with a heteroatom such as, e.g., O, N, or S.
• the heterocycloalkyl optionally has one or more double bonds within a ring, but is not necessarily aromatic.
• the heterocycloalkyl preferably has 3 to about 10 atoms (members) in the carbocyclic skeleton of each ring, preferably from about 4 to about 7 atoms, more preferably from about 5 to about 6 atoms.
• heteroaryl means a radical defined by an aromatic heterocyclic ring as commonly understood in the art, including monocyclic radicals such as, e.g., imidazole, thiazole, pyrazole, pyrrole, furane, pyrazoline, thiophene, oxazole, isoxazole, pyridine, pyridone, pyrimidine, cytosine, 5-methylcytosine, thymine, pyrazine, and triazine radicals, and polycyclics such as, e.g., quinoline, isoquinoline, indole, purine, adenine, guanine, N 6 -methyladenine, and benzothiazole radicals, which heteroaryl radicals are optionally substituted witlrone or more substituents selected from the group consisting of a halogen, an alkyl, alkoxy, an amino, a cyano, a nitro, and the like
• any of the above may be substituted or unsubstitutedwith any suitable substituent.
• suitable substituents include halogen (e.g., fluorine, chlorine, or bromine), alkyl, haloalkyl, alkenyl, alkynyl, alkoxy, haloalkoxy, nitro, cyano, hydroxy, amino, thio, alkylthio, or acyl. These substituents may, themselves, be substituted or unsubstituted.
• the coumarin antibiotic is novobiocin, chlorobiocin, or coumermycin Al, which are shown below.
• the coumarin antibiotic is novobiocin, a well-studied antibiotic whose pharmokinetics and toxicity profile are clearly understood.
• Doses of 4 g/day (well below the maximum tolerated dose) yield a plasma level > 200- 300 ⁇ g/ml, 2 hrs after post-administration, corresponding to a 300-500 ⁇ M drug concentration (see Drusano et al., Antimicrob. Agents Chemother. 30: 42-45 (1986); Eder et al, J. Clin. Invest. 79: 1524-1528 (1987)).
• the contacting of coumarin or a coumarin derivative with the chaperone protein can be carried out in any suitable manner.
• coumarin or a coumarin derivative can be contacted with the chaperone protein by in vivo or ex vivo administration.
• coumarin or a coumarin derivative is administered in vivo to a mammal, more preferably, coumarin or a coumarin derivative is administered in vivo to a human.
• compositions e.g., pharmaceutical compositions
• an acceptable carrier e.g. a pharmacologically acceptable carrier.
• Such compositions can be suitable for delivery of the active ingredient to a patient for medical application, and can be manufactured in a manner that is itself known, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes.
• compositions for use in accordance with the present invention can be formulated in a conventional manner using one or more pharmacologically (e.g., physiologically) acceptable carriers comprising excipients, as well as optional auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically.
• pharmacologically e.g., physiologically
• auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically.
• Proper formulation is dependent upon the route of administration chosen.
• the active ingredient can be formulated in aqueous solutions, preferably in physiologically compatible buffers.
• penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
• the active ingredient can be combined with carriers suitable for inclusion into tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like.
• the active ingredient is conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebuliser, with the use of a suitable propellant.
• the active ingredient can be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion.
• Such compositions can take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and can contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
• Other pharmacological excipients are known in the art.
• the coumarin or coumarin derivative can be administered in unit dosage form, such as a tablet or capsule.
• unit dosage form refers to physically discrete units suitable as unitary dosages for human and animal subjects, each unit containing a predetermined quantity, alone or in combination with other active agents, calculated in an amount sufficient to produce the desired effect in association with a pharmaceutically acceptable diluent, carrier, or vehicle.
• the specifications for the unit dosage forms of the present invention depend on the particular coumarin or coumarin derivative employed and the effect to be achieved, as well as the pharmacodynamics associated with each in the host.
• the dose administered should be an "effective amount” or an amount necessary to achieve an "effective level" in the individual patient.
• the effective level is used as the preferred endpoint for dosing, the actual dose and schedule can vary, depending on interindividual differences in pharmacokinetics, drug distribution, and metabolism.
• One skilled in the art can easily determine the appropriate dose, schedule, and method of administration for the exact formulation of the composition being used, in order to achieve the desired effective level in the individual patient.
• Many of the coumarin or coumarin derivatives are well-know compounds with clearly established effective amounts required to achieve effective levels.
• an appropriate indicator of the effective level of the compounds of the present invention by a direct (e.g., analytical chemical analysis) or indirect analysis of appropriate patient samples (e.g., blood and/or tissues) to determine the suitable dosage required for any given coumarin or coumarin derivative administered.
• the interaction between the chaperone protein and coumarin or a coumarin derivative is such that the chaperone protein does not bind or binds with less affinity to its client protein or client polypeptide.
• Such interference with binding can be accomplished by any suitable method.
• the chaperone protein and coumarin or a coumarin derivative may interact by binding covalently or non- covalently. If non-covalent, the binding can be through hydrogen bonding, ionic bonding, hydrophobic or van der Waals interactions, or any other appropriate type of binding.
• the binding is through non-covalent binding, more preferably, hydrogen or hydrophobic binding.
• a client protein or client polypeptide in the context of the present invention can be any suitable client protein or client polypeptide.
• Client proteins or client polypeptides include, but are not limited to, various transcription factors, including steroid hormone receptors, aryl hydrocarbon receptor, v-ErbA, retinoid receptor, Sim, Myo Dl, Hsf-1, mutated p53, various protein kinases, and various other proteins, such as cytoskeletal proteins, calmodulin, G protein ⁇ -subunits, Proteasome,
• TNF tumor necrosis factor
• retinoblastoma protein Hepatitis B virus reverse transcriptase, tumor necrosis factor (TNF) receptors and retinoblastoma protein. See generally Thormeyer & Baniahmad, Int. J. Mol. Med. 4(4):351-58 (1999) (vErbA); Moffett & Pelletier, FEBS Lett. 466(l):80-6 (2000) (Sim); Jones et al., B. Biol. Sci. 326(1235):277-84 (1990) (Myo Dl); Green et al., Mol. Cell Biol. 15:3354-62 (1995) (Hsf-1).
• steroid hormone receptors include the glucocorticoid receptor, the progesterone receptor, the estrogen receptor, the androgen receptor, and the mineralocorticoid receptor.
• Cytoskeletal proteins include actin, tubulin, and intermediate filaments.
• the client protein or the client polypeptide is a protein kinase. More preferably, the protein kinase is a tyrosine kinase or a serine/threonine kinase.
• Serine/Threonine kinases include the Raf family of kinases, MEK (MAP/ERK (extracellular signal-regulated kinase)), heme regulated ElF-2a kinase, calmodulin dependent eukaryotic elongation factor (eEF)-2 kinase, and casein kinase.
• the client protein or the client polypeptide After binding of the chaperone protein to the coumarin or the coumarin derivative, the client protein or the client polypeptide is generally inactive or less active.
• the client protein or the client polypeptide can be inactive or less active through any suitable method.
• the client protein or the client polypeptide can be degraded by cellular machinery because it is not coupled with its chaperone protein, or it can have an incorrect conformation that distorts its active or recognition site and interferes with the polypeptide performing its designated function.
• the client protein or the client polypeptide can also, e.g., be located in an inappropriate area of the cell, which prevents it from performing its designated function.
• the client protein or client polypeptide is degradation.
• novobiocin a coumarin antibiotic
• a chaperone protein either pure Hsp90, or a solution containing Hsp90 in a cell ly sate, was subsequently incubated with the immobilized novobiocin.
• the cell lysate was preincubated with various members of the coumarin family of antibiotics, namely novobiocin, chlorobiocin, or coumermycin A 1 , and also ATP to determine their ability to inhibit binding of the Hsp90.
• Chlorobiocin and coumermycin A 1 inhibited Hsp90 binding to immobilized novobiocin at about 0.5 mM, while ATP inhibited Hsp90 binding between about 10 and 15 mM, as demonstrated by silver staining.
• novobiocin a coumarin derivative
• Example 2 This example demonstrates in vivo depletion of client protein or client polypeptide through contact of a coumarin or a coumarin derivative with a chaperone protein.
• the level of p60 v src protein was reduced by 50% after exposure to 500 ⁇ M novobiocin.
• Novobiocin also significantly depleted mutant p53 protein in SKBR3 cells, while depleting Raf- 1 protein to an undetectable level in the same cells.
• the steady-state levels of scinderin, an actin-associatedprotein, and BiP (Grp78), an Hsp70 family chaperone protein localized to the ER were not altered by the doses and exposure times of novobiocin used.
• General interference with protein synthesis is also not likely since overnight cyclohexamide treatment of SKBR3 cells did not significantly affect the steady state levels of the proteins affected by novobiocin.
• PBMC peripheral blood mononuclear cells
• the Hsp90 client Raf-1 protein was found to be depleted in a dose-dependent manner by incubation with novobiocin, a coumarin derivative, such that it was almost completely depleted at a concentration of about 0.8 mM novobiocin. Other protein levels remained unaltered, even at the highest drug concentration tested. PBMC remained viable, as assayed by trypan blue.
• C57B16 mice received intraperitoneal injections (100 mg/kg) of novobiocin (a coumarin antibiotic) at 12 hr intervals for 5 days. Animals were sacrificed 3 hrs after the last injection. The spleens were removed, cells were lysed and total protein was assayed by the Bradford method using, e.g., a commercially available kit from BioRad. At this dose, the serum level of novobiocin has been reported to vary between 100 and 450 ⁇ g/ml during the first hr, with the plasma clearance half-life of the drug approximately 80 min. in mice (see Eder at al., Cancer Res. 51 : 510-513 ( 1991 )).
• Raf- 1 levels were determined by Western blotting, as described in Example 2. A control protein, gelsolin, was also assayed using the same method. Optical density of the Raf-1 specific bands was determined using NIH Image software.
• mice receiving novobiocin had significantly lower levels of splenocyte Raf- 1 protein than did controls.
• Raf- 1 protein in the no vobiocin-treated group was reduced by 44% as compared to controls.
• the mean splenocyte Raf-1 protein level was reduced even further, to 29% of control.
• Splenocyte gelsolin remained unchanged in all treated mice.

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• 2000
  • 2000-03-10 EP EP00916277A patent/EP1161231A2/de not_active Withdrawn
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  • 2000-03-10 AU AU37406/00A patent/AU776652B2/en not_active Ceased
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