EP1556392A1 - Molecules de cardiolipine et procedes pour leur synthese - Google Patents

Molecules de cardiolipine et procedes pour leur synthese

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Publication number
EP1556392A1
EP1556392A1 EP03749443A EP03749443A EP1556392A1 EP 1556392 A1 EP1556392 A1 EP 1556392A1 EP 03749443 A EP03749443 A EP 03749443A EP 03749443 A EP03749443 A EP 03749443A EP 1556392 A1 EP1556392 A1 EP 1556392A1
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EP
European Patent Office
Prior art keywords
cardiolipin
composition
acid
liposome
mmol
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.)
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Application number
EP03749443A
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German (de)
English (en)
Inventor
Moghis U. Ahmad
Murali K. Ukkalam
Imran Ahmad
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Neopharm Inc
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Neopharm Inc
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Publication date
Priority claimed from PCT/US2003/013917 external-priority patent/WO2004062569A2/fr
Priority claimed from PCT/US2003/016412 external-priority patent/WO2003099830A2/fr
Application filed by Neopharm Inc filed Critical Neopharm Inc
Publication of EP1556392A1 publication Critical patent/EP1556392A1/fr
Withdrawn legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic Table
    • C07F9/02—Phosphorus compounds
    • C07F9/06—Phosphorus compounds without P—C bonds
    • C07F9/08—Esters of oxyacids of phosphorus
    • C07F9/09—Esters of phosphoric acids
    • C07F9/10—Phosphatides, e.g. lecithin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A61K31/52—Purines, e.g. adenine
    • A61K31/522—Purines, e.g. adenine having oxo groups directly attached to the heterocyclic ring, e.g. hypoxanthine, guanine, acyclovir
    • 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
    • A61K31/7052—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides
    • A61K31/706—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom
    • A61K31/7064—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines
    • A61K31/7068—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines having oxo groups directly attached to the pyrimidine ring, e.g. cytidine, cytidylic acid
    • 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
    • A61K31/7052—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides
    • A61K31/706—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom
    • A61K31/7064—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines
    • A61K31/7068—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines having oxo groups directly attached to the pyrimidine ring, e.g. cytidine, cytidylic acid
    • A61K31/7072—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines having oxo groups directly attached to the pyrimidine ring, e.g. cytidine, cytidylic acid having two oxo groups directly attached to the pyrimidine ring, e.g. uridine, uridylic acid, thymidine, zidovudine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/16—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing nitrogen, e.g. nitro-, nitroso-, azo-compounds, nitriles, cyanates
    • A61K47/18—Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/24—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers
    • 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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers
    • A61K9/1272—Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers with substantial amounts of non-phosphatidyl, i.e. non-acylglycerophosphate, surfactants as bilayer-forming substances, e.g. cationic lipids

Definitions

  • This invention pertains to novel synthetic methods for preparing cardiolipin analogs/variants, and compositions containing them.
  • the invention also pertains to liposome formulations or complexes or emulsions containing active agents or drugs and their use in the treatment of diseases in humans and animals.
  • Liposomal formulations have the capacity to increase the solubility of hydrophobic drugs in aqueous solution. They often reduce the side effects associated with drug therapy and they provide flexible tools for developing new formulations of active agents.
  • Liposomes are commonly prepared from natural phospholipids such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol, phosphatidic acid, and phosphatidylinositol.
  • Anionic phospholipids such as phosphatidyl glycerol and cardiolipin, can be added to generate a net negative surface charge that provides for colloid stabilization. These components are often purified from natural sources and in some cases they can be chemically synthesized.
  • the nature and density of the surface charge of liposomes influences stability, kinetics, biodistribution, and interaction with, and uptake by target cells.
  • Liposome surface charge also influences the tendency for liposomes to aggregate, which makes liposomes difficult to work with and affects uptake by target cells.
  • liposomes with a neutral surface charge have the highest tendency to aggregate, but are less likely to be cleared by cells of the reticuloendothelial system (RES) after systemic administration.
  • RES reticuloendothelial system
  • Negatively charged liposomes exhibit reduced aggregation and increased stability, but exhibit non-specific cellular uptake in vivo.
  • a small amount of negatively charged lipids may stabilize neutral liposomes against an aggregation-dependent uptake mechanism (see, e.g., Drummond et al., Pharm. Rev., 51, 691-743 (1999)).
  • Cardiolipin also known as diphosphatidyl glycerol constitutes a class of complex anionic phospholipids that is typically purified from cell membranes of tissues associated with high metabolic activity, including the mitochondria of heart and skeletal muscles. The negative surface charge of cardiolipin, therefore, stabilizes liposomes against aggregation-dependent uptake, as discussed above.
  • cardiolipin contains up to 90% of linoleic acid (18:2).
  • Yeast cardiolipin differs in having more oleic (18:1) and palmitoleic (16:1) fatty acids, while the bacterial lipid contains saturated and monoenoic fatty acids with 14 to 18 carbons.
  • cardiolipin having shortchain fatty acids are unknown till now.
  • cardiolipin fatty acid chains i.e., saturated or unsaturated
  • Methods for synthesizing cardiolipin comprising short fatty acid chains (“short chain cardiolipin”) have not yet been described.
  • Cardiolipin has also been generated via a reaction between the silver salt of diacylglycerophosphoric acid benzyl ester with 1,3-diiodopropanol benzyl ether or 1,3-diiodopropanol t-butyl ether (see, e.g., De Haas et al., Biochim. Biophys.
  • Phosphate triesters and phosphoramidite esters have been used extensively in nucleic acid synthesis to form phosphate linkages, and to a lesser extent in phospholipid synthesis (see, e.g., Browne et al., J. Chem. Soc. Perkin Trans, 1, 653-657 (2000)). In this respect, Browne et al., supra, describes the preparation of phospholipid analogs, particularly phosphorylcholine analogs, using phosphoramidite methodologies.
  • the phosphatidylinositols PtdIns(4,5)P 2 and PtdIns(3,4,5)P 3 , and derivatives thereof, have been prepared using a variety of phosphoramidite reagents, including N, N- diisopropylphosphoramidite (see, e.g., Watanabe et al., Tetrahedron Lett. 35, 123-124 (1994)), difluorenyl phosphoramidite (see, e.g., Watanabe et al., Tetrahedron Lett.
  • phosphotriester analogs of PtdIns(4,5)P 2 and PtdIns(3,4,5)P 3 have been prepared utilizing the phosphoramidite reagent 2-cyano-ethyl N, N, N, N- tetraisopropylphosphorodiamidite (see, e.g., Gu et al, J. Org. Chem, 61, 8642-8647 (1996)).
  • Chem, 64, 648-651 (1999) describe the synthesis of phosphatidyl glycerol from 2,5-diben2yl-D-mannitol utilizing methyl tetraisopropylphosphorodiamidite as a phosphorylating agent.
  • phosphate triesters and phosphoramidite esters in preparing phospholipids such as cardiolipin, particularly cardiolipin species having varying fatty acid chain lengths, however, is not well established.
  • the invention provides novel synthetic methodologies for preparing cardiolipin having varying fatty acids and/or alkyl chains with varying length and saturation/unsaturation.
  • the methods comprises of (a) reacting an optically pure 1,2-O- diacyl-OT-glycerol or l,2-O-dialkyl-jn-glycerol with one or more phosphoramidite reagent(s) or one or more phosphate triester(s), (b) coupling the product of (a) with a 2- protected glycerol, wherein a protected cardiolipin is produced, and (c) deprotecting the protected cardiolipin, such that the cardiolipin is prepared.
  • the invention also provides a method for preparing cardiolipin having varying fatty acid chain lengths comprising (a) reacting a 2-O-protected glycerol with one or more phosphoramidite reagents, wherein a phosphorylating agent is produced, (b) reacting the phosphorylating agent with an optically pure 1,2-O-diacyl-sr ⁇ -glycerol or l,2-O-dialkyl-,sn-glycerol, wherein a protected cardiolipin is produced, and (c) deprotecting the protected cardiolipin, such that the cardiolipin is prepared.
  • the cardiolipin prepared by the present methods can be incorporated into liposomes, which can also include active agents such as hydrophobic or hydrophilic drugs, antisense nucleotides or diagnostic agents. Such liposomes can be used to treat diseases or in diagnostic and/or analytical assays.
  • Figure 1 depicts the general structure of cardiolipin.
  • Figure 2 depicts one scheme for synthesizing cardiolipin.
  • Figure 3 depicts an alternative synthetic scheme for cardiolipin.
  • Figure 4 depicts an alternative synthetic scheme for cardiolipin.
  • Figure 5 depicts an alternative synthetic scheme for cardiolipin.
  • Figure 6 depicts an alternative synthetic scheme for cardiolipin ether analogs.
  • Figure 7 depicts an alternative synthetic scheme for cardiolipin ether analogs.
  • the present invention describes methods for the synthesis of cardiolipin variants and analogs having the general formulas I, II, and III, as well as compositions containing such variants and analogs.
  • Yj. andY 2 are the same or different and are -O-C(O)-, -O-, -S-, -NH-C(O)- or the like.
  • R t and R 2 are the same or different and are H, saturated and/or unsaturated alkyl group, preferably a C 2 to C 34 saturated and/or unsaturated alkyl group.
  • R 4 is hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, a peptide, dipeptide, polypeptide, protein, carbohydrate (such as glucose, mannose, galactose, polysaccharide and the like), lieterocyclic, nucleoside, polynucleotide and the like.
  • R 5 is a linker, which may (or may not be) added in the molecule depending on the need and applications.
  • R 5 can comprise alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, alkoxy, polyalkyloxy (such as pegylated ether of containing from about 1 to 500 alkyloxymers (and can have at least about 10 alkyloxy mers, such as at least about 50 alkyloxy mers or at least about 100 alkyloxy mers, such as at least about 200 alkyloxy mers or at least about 300 alkyloxy mers or at least about 400 alkyloxy mers), substituted polyalkyloxy and the like), a peptide, dipeptide, polypeptide, protein, carbohydrate such as glucose, mannose, galactose, polysaccharides and the like.
  • X is hydrogen or a non-toxic cation, preferably ammonium, sodium, potassium, calcium, barium ion and the like.
  • alkyl encompasses saturated or unsaturated straight-chain and branched-chain hydrocarbon moieties.
  • substituted alkyl comprises alkyl groups further bearing one or more substituents selected from hydroxy, alkoxy (of a lower alkyl group), mercapto (of a lower alkyl group), cycloalkyl, substituted cycloalkyl, halogen, cyano, nitro, amino, a ido, imino, thio, -C(O)H, acyl, oxyacyl, carboxyl, and the like.
  • Y ⁇ and Y 2 in Formula III are -O-C(O)- or -O-.
  • R 3 is most preferably is CH 2 .
  • Ri and R 2 are the same and are a C 2 to C 13 saturated and/or unsaturated alkyl group, more preferably between 4 and 14 carbon atoms (such as between about 6 and 12 carbon atoms).
  • X is most preferably is hydrogen or ammonium ion. In the absence of linker (R 5 ), it gives the general structure of cardiolipin ( Figure 1).
  • the invention provides a method for preparing cardiolipin or an analogue thereof of Formulas I, II, or III, comprising reacting an alcohol of the formula VIII with one or more phosphoramide reagents and 2-O-protected glycerol or 2-O-subsituted glycerol VI in the presence of an acid catalyst.
  • Y in formula VI is a hydroxyl protecting group, preferably alkyl group or the like, or a silyl protecting group.
  • Ri, R 2 , R 3 , Yi, and Y 2 can be as indicated above with respect to Formulas I, II, or ⁇ i.
  • the acid catalyst can be any suitable catalyst that can facilitate the reaction.
  • catalysts examples include 4,5-dichloroimidazole, lH-tetrazole, 5-(4-nitrophenyl)-lH-tetrazole, 5-(3,5- dinitrophenyl)-lH-tetrazole, N-methylimidazolium triflate, and N-methylimidazolium perchlorate, 4,5-dicyanoimidazole, 5-ethylthio-lH-tetrazole, and 5-methylthio-lH-tetrazole.
  • Preferred catalysts are 4,5-dichloroimidazole orlH-tetrazole.
  • the coupling phosphoramidites can have formula IV or V:
  • the invention provides a method for preparing cardiolipin or an analogue thereof of formulas I, II, or III; comprising reacting 2-O protected glycerol with one or more phosphotriesters in the presence of pyridinium tribromide.
  • Preferred phosphotriesters can be produced by reacting an alcohol of formula VIII with phosphoramidite of general formula VII.
  • X in Formulas IV, V, or VII is a phosphate protecting group, preferably a benzyl group or 2-cyanoethyl or silyl group.
  • suitable protecting groups include alkyl phosphates including ethyl, cyclohexyl, t-butyl; 2-substituted ethyl phosphates including 2-cyanoethyl, 4-cyano-2-butenyl, 2-(methyldiphenylsilyl)ethyl, 2- (trimethylsily ⁇ )ethyl, 2-(triphenylsilyl)ethyl; haloethyl phosphates including 2,2,2- trichloroethyl, 2,2,2-tribromoethyl, 2,2,2-trifluoroethyl; benzyl phosphates including 4- chlorobenzyl, fluorenyl-9-methyl, diphenylmethyl and amidates.
  • FIG. 2 & 3 A general sequence of reactions for the synthesis of compound of invention is illustrated in Figures 2 & 3.
  • the present invention provides a method for preparing cardiolipin I having varying fatty acid chain lengths comprising (a) reacting an optically pure l,2-O-diacyl--f «-glycerol 2 with one or more phosphoramidite reagent(s)of the general formula IV (figure 2) or V (figure 3) (b) coupling the product of (a) 3 with a 2-O-protected glycerol VI in a chlorinated solvent (for example dichloromethane, chloroform or like) followed by oxidation with m-chloroperoxybenzoic acid (m-CPBA) results in the production of a protected cardiolipin 4. Thereafter, deprotecting the protected cardiolipin followed by conversion to ammonium salt will result in the production of cardiolipin 1 (ammonium salt).
  • a chlorinated solvent for example dichloromethane, chloroform
  • any suitable phosphoramidite reagent or methodology may be used, such as is described in, for example Browne et al., supra.
  • suitable phosphoramidite reagents include (benzyloxyXN N-diisopropylamino)chlorophosphine (see, e.g., Prestwich et al. J. Am. Chem. Soc. 1991, 113, 1822-1825), benzyloxybis (diisopropylamino) phosphine (see, e.g., Dreef et al. Tetrahedron Lett.
  • the optically pure l,2-O-diacyl-,s72-glyeerol 2 can be phosphorylated using phosphoramidite VII to yield phosphite triesters 5 which can be coupled with any suitable 2-O-protected glycerol VI, such as, for example, benzyloxy 1,3-propanediol or 2- levulinoyl-1.
  • 2-O-protected glycerol VI such as, for example, benzyloxy 1,3-propanediol or 2- levulinoyl-1.
  • 3-propanediol using pyridinium perbromide and phosphoniurn salt methodology (see, e.g., Watanabe et al., supra) to get protected cardiolipin 4.
  • the preferred coupling reagent in this context of synthetic methods is dibenzyl diisopropylphosphoramidite.
  • the inventive method comprises (a) reacting a 2-O-protected glycerol VI with one or more phosphoramidite reagents IV or V, wherein a phosphorylating agent 6 is produced, (b) reacting the phosphorylating agent 6 with an optically pure l,2-O-diacyl-s «-glycerol 2 followed by oxidation with m-CPBA, wherein a protected cardiolipin 4 is produced, and (c) deprotecting the protected cardiolipin, such that the cardiolipin is prepared.
  • Suitable phosphoramidite reagents and 2-O-protected glycerols for use in this aspect of the inventive method are described above.
  • FIG. 6 Another embodiment of the present invention, represented in Figure 6 leads to ether analogs of cardiolipin, wherein the acyl groups are replaced by alkyl chain. Accordingly (a) 1,2-O-dialkyl- ⁇ -glycerol 7 is treated with phosphoramidites IV or V wherein a phosphorylating agent 8 is produced, (b) reacting the phosphorylating agent with a 2-O-protected glycerol VI followed by oxidation, wherein a protected cardiolipin 9 is produced, and (c) deprotecting the protected cardiolipin, such that the ether analog of cardiolipin 10 is produced. [0030] Another embodiment of the present invention is depicted in Figure 7.
  • the optically pure 1,2-O-dialkyl-OT-glycerol 7 can be phosphorylated using phosphoramidite VII to yield phosphite triesters 11 which can be coupled with any suitable 2-O-protected glycerol VI, such as, for example, benzyloxy 1,3-propanediol or 2- levulinoyl-1, 3-propanediol using pyridinium perbromide and phosphonium salt methodology (see, e.g., Watanabe et al., supra) to get protected cardiolipin ether analog 9.
  • the preferred coupling reagent in this context of synthetic methods is dibenzyl diisopropylphosphoramidite.
  • the invention described above is an elegant and efficient method of synthesizing cardiolipin.
  • the routes are short and proceed in good overall yield.
  • the deprotection can be accomplished by a method depending on the protecting group.
  • a benzyl group can be removed by catalytic hydrogenolysis or by treatment with Nal, 2-cyanoethyl and fluorenylmethyl groups by treatment with a tertiary base such as triethylamine
  • a silyl group can be deprotected with fluoride ion or acidic medium, a levulinoyl group by hydrazinolysis.
  • phosphoramidites and phosphate esters can be prepared using a variety of acid catalysts, including 4,5-dichloroimidazole (see, e.g., Browne et al.), 5-(4-nitrophenyl)-lH- tetrazole, 5-(3,5-dinitrophenyl)-lH-tetrazole, N-methylimidazolium triflate, and N- methylimidazolium perchlorate (see, e.g., Moriguchi et al.).
  • tert- butylhydroperoxide can be used as an alternative oxidant.
  • the described methods can be further modified in any suitable manner known in the art.
  • the inventive method can be used to prepare cardiolipin species comprising fatty acid /alkyl chains of varying length and saturation/unsaturation.
  • the general structure of a phospholipid fatty acid comprises a hydrocarbon chain and a carboxylic acid group.
  • the length of the fatty acid hydrocarbon chain ranges from about 2 to about 34 carbon atoms and can be saturated or unsaturated.
  • the carbon chain is more typically between about 12 and about 24 carbon atoms.
  • the length of the fatty acid hydrocarbon is less than about 24 carbon acids, such as less than about 24 carbon atoms, or even less than about 20 carbon atoms.
  • the invention also provides a cardiolipin or cardiolipin analogue prepared in accordance with the inventive method.
  • the cardiolipin prepared by the inventive method comprises a short fatty acid chain (i.e., a "short chain cardiolipin"), and the invention provides a short chain cardiolipin.
  • a short fatty acid chain comprises between about 2 and between about 14 carbon atoms, and can have between about 4 (or about 6) and about 12 carbon atoms, such as between about 8 and about 10 carbon atoms.
  • the cardiolipin produced by the inventive method can comprise a long chain fatty acid chain (i.e., a "long chain cardiolipin").
  • a long fatty acid chain comprises between about 14 and about 34 carbon atoms, such as between about 14 (or between about 20) and about 24 carbon atoms.
  • the inventive method is not limited to the production of short or long chain cardiolipin species exclusively. Indeed, a cardiolipin containing fatty acid alkyl chains of intermediate length can also be prepared by the inventive method.
  • Phospholipid fatty acids typically are classified by the number of double and/or triple bonds in the hydrocarbon chain (i.e., unsaturation).
  • a saturated fatty acid does not contain any double or triple bonds, and each carbon in the chain is bound to the maximum number of hydrogen atoms.
  • the degree of unsaturation of a fatty acid depends on the number of double or triple bonds present in the hydrocarbon chain. In this respect, a mon ⁇ unsaturated fatty acid contains one double bond, whereas a polyunsaturated fatty acid contains two or more double bonds (see, e.g., Oxford Dictionary of Biochemistry and Molecular Biology, rev. ed., A.D. Smith (ed.), Oxford University Press (2000), and Molecular Biology of the Cell, 3 rd ed., B.A. Alberts (ed.), Garland Publishing, New York (1994)).
  • the fatty acid chains of the cardiolipin are prepared by the inventive method, whether short or long, also can be saturated or unsaturated.
  • the described methods can be used to prepare a variety of novel cardiolipin molecules.
  • the methods can be used to prepare cardiolipin variants in pure form containing short or long fatty acid side chains.
  • Preferred fatty acids range from carbon chain lengths of about C 2 to C 3 , preferably between about C and about C 2 , and include tetranoic acid (C 4: o), pentanoic acid (C 5 :o), hexanoic acid (C ⁇ -.o heptanoic acid (C 7: o), octanoic acid (C 8: o), nonanoic acid (C 9: o), decanoic acid (C ⁇ o : o), undecanoic acid (C 11: o), dodecanoic acid (C ⁇ 2: o), tridecanoic acid (C 13: o), tetradecanoic (myristic) acid (C ⁇ : o), pentadecanoic acid (Ci5 : o), pen
  • the alkyl chain will also range from C 2 to C 3 preferably between about C and about C 2 .
  • Other fatty acid chains also can be employed as Ri and or R 2 substituents. Examples of such include saturated fatty acids such as ethanoic (or acetic) acid, propanoic (or propionic) acid, butanoic (or butyric) acid, hexacosanoic (or cerotic) acid, octacosanoic (or montanic) acid, triacontanoic (or melissic) acid, dotriacontanoic (or lacceroic) acid, tetratriacontanoic (or gheddic) acid, pentatriacontanoic (or ceroplastic) acid, and the like; monoefhenoic unsaturated fatty acids such as trcms ⁇ -butenoic (or crotonic) acid, cis-2- butenoic (
  • 'hydroxyl protecting group' used herein the invention refers to the commonly used protecting groups disclosed by T. W. Greene and P. G. Wuts, Protective Groups in Organic Synthesis, 3 rd edition, John Wiley & Sons, New York (1999).
  • Such protecting groups include methyl ether, substituted methyl ethers including methoxymethyl, benzyloxymethyl, ⁇ -methoxybenzyloxymethyl, 2-methoxyethoxymethyl, tetrahydropyranyl, tetrahydrofuranyl ethers; substituted ethyl ethers like 1-ethoxyethyl, 1- methyl-1-benzyloxy ethyl, allyl, propargyl; benzyl and substituted benzyl ethers including p- methoxybenzyl, 3,4-dimethoxybenzyl, triphenylmethyl; silyl ethers including trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, diphenylmethylsilyl; esters including formate, acetate, chloroacetate, dichloroacetate, trichloroacetate, benzoate, levuliny
  • 'phosphate protecting group' used herein the invention refers to the commonly used protecting groups described by T. W. Greene and P. G. Wuts, Protective Groups in Organic Synthesis, 3 rd edition, John Wiley & Sons, New York (1999).
  • Such protecting groups include alkyl phosphates including methyl, ethyl, cyclohexyl, t-butyl; 2- substituted ethyl phosphates including 2-cyanoethyl, 4-cyano-2-butenyl, 2- (methyldiphenylsilyl)ethyl, 2-(trimethylsilyl)ethyl, 2-(triphenylsilyl)ethyl; haloethyl phosphates including 2,2,2-trichloroethyl, 2,2,2-tiibromoethyl, 2,2,2-trifluoroethyl; benzyl phosphates including 4-chlorobenzyl, fluorenyl-9-methyl, diphenylmethyl and amidates.
  • the cardiolipin molecules described herein and cardiolipins produced by the inventive method can be used in lipid formulations, such as liposomal compositions. Complexes, emulsions and other formulations including the inventive cardiolipin also are within the scope of the present invention.
  • Such formulations according to the present invention can be prepared by any suitable technique.
  • the invention provides a method for preparing a liposome or other lipid composition, comprising preparing a cardiolipin or cardiolipin analogue as described herein and including the cardiolipin or cardiolipin analogue in a lipid formulation, such as a liposome.
  • the invention also includes such lipid compositions including the inventive cardiolipin and/or cardiolipin analogues.
  • the liposomal composition, complex, emulsion and the like can include other lipids.
  • the composition can include one or more phosphatidylcholines, such as, for example, dimyristoylphosphatidylcholine, distearoylphosphatidylcholine, dioleylphosphatidylcholine, dipalmitoylphosphatidylcholine, diarachidonoylphosphatidylcholine, egg phosphatidylcholine, soy phosphatidylcholine, hydrogenated soy phosphatidylcholine, and mixtures thereof.
  • phosphatidylcholines such as, for example, dimyristoylphosphatidylcholine, distearoylphosphatidylcholine, dioleylphosphatidylcholine, dipalmitoylphosphatidylcholine, diarachidonoylphosphatidylcholine, egg phosphatidylcholine, soy phosphatidyl
  • the composition can include one or more phosphatidylglycerols, such as dimyristoylphosphatidylglycerol, distearoylphosphatidylglycerol, dioleylphosphatidylglycerol, dipalmitoylphosphatidylglycerol, diarachidonoylphosphatidylglycerol, and mixtures thereof.
  • the composition can include one or more sterols, such as cholesterol, derivatives of cholesterol, coprostanol, cholestanol, cholestane, cholesterol hemisuccinate, cholesterol sulfate, and mixtures thereof.
  • the composition in addition to the cardiolipin or cardiolipin analogue, includes a phosphatidylcholine, a sterol, and a tocopherol (e.g., ⁇ -tocopherol).
  • the composition also can include stabilizers, absorption enhancers, antioxidants, phospholipids, biodegradable polymers and medicinally active agents among other ingredients.
  • it is preferable for the inventive composition, especially liposomal composition to include one or more targeting agents, such as carbohydrate or a protein or other ligand that binds to a specific substrate, for example, that recognize cellular receptors.
  • the composition also can include one or more active agents.
  • a single active agent can be included, or a mixture of active agents (e.g., two or more active agents) can be included within the composition.
  • Active agents can be present in any suitable manner in the composition. For example, they can be complexed with the cardiolipin or cardiolipin analogue in the composition. Additionally, or alternatively, one or more active agents can be entrapped within liposomes, when the composition is a liposomal composition.
  • Active agents which are compatible with the present invention include, for example, agents which act on the peripheral nerves, adrenergic receptors, cholinergic receptors, the skeletal muscles, the cardiovascular system, smooth muscles, the blood circulatory system, synaptic sites, neuroeffector junctional sites, endocrine and hormone systems, the immunological system, the reproductive system, the skeletal system, the alimentary and excretory systems, the histamine system and the central nervous system.
  • Suitable agents may be selected from, for example, proteins, enzymes, hormones, nucleotides (including sense and antisense oligonucleotides (see, e.g., U.S.
  • Patent 6,126,965 polynucleotides, nucleoproteins, polysaccharides, glycoproteins, lipoproteins, polypeptides, steroids.
  • Active agents can be analgesics, anesthetics, anti-arrythmic agents, antibiotics, antiallergic agents, antifungal agents, anticancer agents, anticoagulants, antidepressants, antidiabetic agents, anti-epilepsy agents, anti-inflammatory corticosteroids, agents for treating Alzheimers or Parkinson's disease, antiulcer agents, anti-protozoal agents, anxiolytics, thyroids, anti-thyroids, antivirals, anoretics, bisphosphonates, cardiac inotropic agents, cardiovascular agents, corticosteroids, diuretics, dopaminergic agents, gastrointestinal agents, hemostatics, hypercholesterol agents, antihypertensive agents (e.g., dihydropyridines), antidepressants, and cox-2 inhibitors, immunosuppressive agents, anti- g
  • the therapeutic agents can be nephrotoxic, such as cyclosporins and amphotericin B, or cardiotoxic, such as amphotericin B and paclitaxel.
  • exemplary anticancer agents include melphalan, chlormethine, extramustinephosphate, uramustine, ifosfamide, mannomustine, trifosfamide, streptozotocin, mitobronitol, mitoxantrone (see., e.g., published international patent application WO 02/32400), methotrexate, fluorouracil, cytarabine, tegafur, idoxide, taxanes (e.g., taxol, paclitaxel, etc., see published international patent application WO 00/01366), daunomycin, daunorubicin, bleomycin, amphotericin, carboplatin, cisplatin, paclitaxel, BC ⁇ U, vinca alkaloids (e.
  • drugs which may be delivered according to the method include, prochlorperzine edisylate, ferrous sulfate, aminocaproic acid, mecamylamine hydrochloride, procainamide hydrochloride, amphetamine sulfate, methamphetamine hydrochloride, benzamphetamine hydrochloride, isoproterenol sulfate, phenmetrazine hydrochloride, bethanechol chloride, methacholine chloride, pilocarpine hydrochloride, atropine sulfate, scopolamine bromide, isopropamide iodide, tridihexethyl chloride, phenformin hydrochloride, metliylphenidate hydrochloride, theophylline cholinate, cephalexin hydrochloride, diphenidol, meclizine hydrochloride, prochlorperazine maleate, phenoxybenzamine, thiethy
  • proteins and peptides which include, but are not limited to, bone morphogenic proteins, insulin, colchicine, glucagon, thyroid stimulating hormone, parathyroid and pituitary hormones, digestive hormones, calcitonin, renin, prolactin, corticotrophin, thyrotropic hormone, follicle stimulating hormone, chorionic gonadotropin, gonadotropin releasing hormone, bovine somatotropin, porcine somatotropin, oxytocin, vasopressin, GRF, somatostatin, lypressin, pancreozymin, luteinizing hormone, LHRH, LHRH agonists and antagonists, leuprolide, interferons (e.g., consensus interferon, interferon ⁇ -2a, interferon cc-2b, ⁇ -, ⁇ -, or ⁇ - interferons), interleuldns, growth hormones such as human growth hormone and its derivatives such as methione-human
  • liposomes can have a net neutral, negative or positive charge.
  • positive liposomes can be formed from a solution containing phosphatidylcholine, cholesterol, cardiolipin and enough stearylamine to overcome the net negative charge of cardiolipin.
  • Negative liposomes can be formed from solutions containing phosphatidyl choline, cholesterol, and/or cardiolipin variants prepared by the methods described herein.
  • the liposomes of the present invention can be multi or unilamellar vesicles depending on the particular composition and procedure to make them.
  • Liposomes can be prepared to have substantially homogeneous sizes in a selected size range, such as about 1 micron or less, or about 500 nm or less, about 200nm or less, or about lOOnm or less.
  • One effective sizing method involves extruding an aqueous suspension of the liposomes through a series of polycarbonate membranes having a selected uniform pore size; the pore size of the membrane will correspond roughly with the largest sizes of liposomes produced by extrusion through that membrane.
  • the liposomal (or other lipid) composition can be in any desired form.
  • the composition can be ready for administration to a patient.
  • the composition can be in dried or lyophilized form.
  • cryoprotectant include, for example, sugars such as tiehalose, maltose, lactose, sucrose, glucose, and dextran, with the most preferred sugars from a performance point of view being tiehalose and sucrose.
  • sugars such as tiehalose, maltose, lactose, sucrose, glucose, and dextran
  • Other more complicated sugars can also be used, such as, for example, aminoglycosides, including streptomycin and dihydrostreptomycin.
  • Any suitable method can be employed to form the liposomes.
  • lipophilic liposome-forming ingredients such as phosphatidylcholine, a cardiolipin prepared by the methods described above, cholesterol and ⁇ -tocopherol can be dissolved or dispersed in a suitable solvent or combination of solvents and dried.
  • suitable solvents include any non-polar or slightly polar solvent, such as t-butanol, ethanol, methanol, chloroform, or acetone that can be evaporated without leaving a pharmaceutically unacceptable residue. Drying can be by any suitable means such as by lyophilization. The dehydration is typically achieved under vacuum and can take place either with or without prior freezing of the liposome preparation.
  • Hydrophilic ingredients can be dissolved in polar solvents, including water.
  • the invention provides a method for retaining a drug in a liposome.
  • cardiolipin or cardiolipin analogue is prepared as described herein, and the cardiolipin or cardiolipin analogue and a drug or drugs (e.g., an active agent a mixture of active agents) is included within a liposome.
  • active agent(s) can be dissolved or dispersed in a suitable solvent and added to the liposome mixture prior to mixing.
  • hydrophilic active agents will be added directly to the polar solvent and hydrophobic active agents will be added to the nonpolar solvent used to dissolve the other ingredients but this is not required.
  • the active agent could be dissolved in a third solvent or solvent mix and added to the mixture of polar solvent with the lipid film prior to homogenizing the mixture.
  • Liposomes can be coated with a biodegradable polymers such as sucrose, epichlorohydrin, branched hydrophilic polymers of sucrose, polyethylene glycols, polyvinyl alcohols, methoxypolyethylene glycol, ethoxypolyethylene glycol, polyethylene oxide, polyoxyethylene, polyoxypropylene, cellulose acetate, sodium alginate, ⁇ , ⁇ - diefhylaminoacetate, block copolymers of polyoxyethylene and polyoxypropylene, polyvinyl pyrrolidone, polyoxyethylene X-lauryl ether wherein X is from 9 to 20, and polyoxyethylene sorbitan esters.
  • a biodegradable polymers such as sucrose, epichlorohydrin, branched hydrophilic polymers of sucrose, polyethylene glycols, polyvinyl alcohols, methoxypolyethylene glycol, ethoxypolyethylene glycol, polyethylene oxide, polyoxyethylene, polyoxypropylene, cellulose a
  • Antioxidants can be included in the liposomal composition or other lipid composition. Suitable antioxidants include compounds such as ascorbic acid, tocopherol, and deteroxime mesylate.
  • Absorption enhancers can be included in the liposomal composition or other lipid composition. Suitable absorption enhancers include Na-salicylate-chenodeoxy cholate, Na deoxycholate, polyoxyethylene 9-lauryl ether, chenodeoxy cholate- deoxycholate and polyoxyethylene 9-lauryl ether, monoolein, Na tauro-24,25- dihydrofusidate, Na taurodeoxycholate, Na glycochenodeoxycholate, oleic acid, linoleic acid, linolenic acid.
  • inventive lipid (e.g., liposomal) composition also can include one or more pharmaceutically acceptably excipients.
  • pharmaceutically suitable excipients include solid, semi-solid or liquid diluents, fillers and formulation auxiliaries of all kinds.
  • the invention also includes pharmaceutical preparations in dosage units.
  • the preparations are in the form of individual parts, for example vials, syringes, capsules, pills, suppositories, or ampoules, of which the content of the liposome formulation of active agent corresponds to a fraction or a multiple of an individual dose.
  • the dosage units can contain, for example, 1, 2, 3, or 4 individual doses, or 1/2, 1/3, or 1/4 of an individual dose.
  • An individual dose preferably contains the amount of active agent which is given in one administration and which usually corresponds to a whole, a half, a third, or a quarter of a daily dose.
  • Tablets, dragees, capsules, pills, granules, suppositories, solutions, suspensions and emulsions, pastes, ointments, gels, creams, lotions, powders and sprays can be suitable pharmaceutical preparations.
  • Suppositories can contain, in addition to the liposomal active agent, suitable water-soluble or water-insoluble excipients. Suitable excipients are those in which the inventive liposomal active agent is sufficiently stable to allow for therapeutic use, for example polyethylene glycols, certain fats, and esters or mixtures of these substances.
  • Ointments, pastes, cream, and gels can also contain suitable excipients in which the liposomal active agent is stable.
  • the composition also can be formulated for injection (e.g., intravenously, interstitially, intratumorally, etc) by the inclusion of one or more excipients (e.g., buffered saline) suitable for injection.
  • the active agent or its pharmaceutical preparations can be administered intravenously, subcutaneously, locally, orally, parenterally, intraperitoneally, and/or rectally or by direct injection into tumors or sites in need of treatment by such methods as are known or developed.
  • Cardiolipin and cardiolipin-analog based formulations also can be administered topically, e.g., as a cream, skin ointment, dry skin softener, moisturizer, etc.
  • the invention provides for the use of the composition to prepare a medicament for the treatment of a disease. In this sense, the invention also provides a method for treating a human or animal disease.
  • the inventive composition is exposed to (administered to) a human or animal patient in need of such treatment.
  • the inventive method facilitates delivery of the active agent(s) to the patient.
  • the method can be used to administer one or more active agents. It is thought to be general for active agents that are stable in the presence of surfactants. Hydrophilic active agents are suitable and can be included in the interior of the liposomes such that the liposome bilayer creates a diffusion barrier preventing it from randomly diffusing throughout the body. Hydrophobic active agents are thought to be particularly well suited for use in the present method because they not only benefit by exhibiting reduced toxicity but they tend to be well solubilized in the lipid bilayer of liposomes.
  • Suitable diseases for treatment will depend on the selection of active agents, such as described herein. However a preferred disease is cancer, in which instance, at least one active agent incorporated into the composition is an anticancer agent.
  • Chemotherapeutic agents are well suited for such use. Liposome formulations containing chemotherapeutic agents may be injected directly into the tumor tissue for delivery of the chemotherapeutic agent directly to cancer cells. In some cases, particularly after resection of a tumor, the liposome formulation can be implanted directly into the resulting cavity or may be applied to the remaining tissue as a coating.
  • the method can be employed to treat diseases, disorders, or symptoms within patients even where the composition does not contain an active pharmaceutical agent other than cardiolipin.
  • the invention provides for the use of cardiolipin to prepare a medicament to combat or treat such diseases, disorders, or symptoms.
  • the invention further provides a method of treating such diseases, disorders, or symptoms within patients, and the effects of such diseases, disorders, or symptoms by administering to the patient a therapeutically effective amount of cardiolipin.
  • cardiolipin provides a beneficial antioxidant effect, which can alleviate the effects of many diseases, disorders, or symptoms.
  • conditions that can be treated in accordance with the method include, for example, age-related diseases, atherosclerosis, diabetes, heart disease, ischemia, and skin disorders (e.g., acne, psoriasis, eczema, etc.).
  • the method also can be employed to combat the effects of aging.
  • the cardiolipin can be formulated as a liposomal or non- liposomal formulation (e.g., an emulsion, cream, etc.) as discussed herein and can include, in addition to cardiolipin, one or more pharmaceutically acceptable carriers.
  • the composition can be administered by any suitable route.
  • the composition can be administered dermally, intravenously, or by other desired route of administration.
  • the invention also is directed to methods of delivering active agents (or mixtures of active agents) to cells.
  • the methods can be carried out by preparing liposomes that include active agents and cardiolipin variants/analogs as synthesized by the above disclosed methods.
  • the liposomes are then delivered to a cell or cells, which can be in vitro or in vivo, as desired. In vivo administration can be achieved as described herein or as otherwise known to those of ordinary skill.
  • composition e.g., liposomes
  • delivery of the active agent(s) can be carried out by adding the composition (e.g., liposomes) to the cell culture medium, for example.
  • Ri, R 2 myristoyl (C 1 : o chain)
  • Ri, R2 myristoyl (C 14:0 chain)
  • the catalyst was filtered off over celite bed, treated with 4 mL of 30% ammonia solution and concentrated, the residue was dissolved in CHC1 3 , filtered through a 0.25 ⁇ filter and precipitated with acetone to give tetramyristoyl (Cj-j.-o) cardiolipin (1.75 g, 83%) as a white solid.
  • R l5 R 2 lauroyl (C ⁇ 2: o chain)
  • Method 1 A solution of 1 ,2-dilauroyl-- -glycerol (2.2 g, 4.82 mmol), benzyl N N-tetraisopropyl phosphoramidite (1.95 g, 5.78 mmol) and lH-tetrazole (12.84 mL of 0.45 M sol in acetonitrile, 5.78 mmol) in CH 2 C1 2 (25 mL) was stirred at room temperature under argon for 3 h.
  • Method 2 To a stirred solution of 1,2-Dilauroyl-r ⁇ -glycerol (5.0 g, 10.96 mmol) and tetrazole (29.2 mL of 0.45 M sol in acetonitrile, 13.15 mmol) in 40 mL anhydrous CH 2 CI 2 , dibenzyl diisopropyl phosphoramidite (4.54g, 13.15 mmol) was added and stirred at room temperature for 2 h.
  • Ri, R 2 lauroyl (C 12: o chain)
  • Ri, R2 lauroyl (C 12: o chain)
  • Ri, R 2 decanoyl ( oio chain)
  • Ri, R2 decanoyl (C ⁇ o chain)
  • Ri, R 2 decanoyl (C ⁇ o : o chain)
  • Ri, R2 octanoyl (C 8: o chain)
  • Ri, R2 octanoyl (C 8: o chain)
  • R l5 R 2 octanoyl (C 8: o chain)
  • l,2-Dioctanoyl-,s' «-glycerol from 5B (5.0 g, 14.53 mmol) and tetrazole (40.3 mL of 0.45 M sol in acetonitrile, 18.16 mmol) in 50 mL anhydrous CH 2 C1 2 dibenzyl diisopropyl phosphoramidite (6.26g, 18.16 mmol) was added and stirred at room temperature for 2 h.
  • Ri, R2 hexanoyl (C 6: o chain)
  • Ri, R2 oleoyl (C 18:1 chain)
  • R ls R 2 hexyl (C 6: o chain)
  • Example 10 This example demonstrates preparation of a cardiolipin-containing liposome composition of the invention.
  • Small unilamellar vesicles are formed by mixing 19.1 ⁇ mole of cardiolipin, produced according to the methods described herein, 96.2 ⁇ mol of phosphatidyl choline and 64.6 ⁇ mol of cholesterol. After thorough stirring, the mixture is evaporated to dryness in a 50 ml round-bottom flask using a rotary evaporator. The subsequent dried lipid film is resuspended in 10 ml sterile non-pyrogenic water. After a 30 minute swelling time, the resulting suspension is sonicated in a fixed temperature bath at 25 °C for 15 minutes. The preparation of liposomes is then lyophilized with tiehalose.

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Abstract

La présente invention concerne de nouvelles voies de la synthèse de la cardiolipine avec différents acides gras et/ou chaînes alkyle de longueur de chaîne variable, avec ou sans insaturation, et plus particulièrement une cardiolipine à chaîne courte. Pour ce procédé, on prend un 1,2-O-sn-diacyle/1,2-O-sn-dialkyle glycérol ou un glycérol 2-O-protégé, et on le fait réagir avec un réactif au phosphoramidite ou un triester de phosphate de façon à produire une cardiolipine protégée que l'on déprotège pour obtenir la cardiolipine à chaîne courte. Ces logiques de réaction se prêtent à l'obtention de nouvelles variantes de cardiolipine. La cardiolipine ainsi obtenue peut s'incorporer à des liposomes pouvant également inclure des agents actifs tels que des médicaments hydrophobes ou hydrophiles. De tels liposomes conviennent pour le traitement de maladies ou pour des essais de diagnostic ou d'analyse. Les liposomes peuvent également comporter des ligands permettant de cibler un type de cellule particulier ou un tissu spécifique.
EP03749443A 2002-10-16 2003-09-05 Molecules de cardiolipine et procedes pour leur synthese Withdrawn EP1556392A1 (fr)

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US438659P 2003-01-07
US46733103P 2003-05-02 2003-05-02
US467331P 2003-05-02
WOPCT/US03/13917 2003-05-04
PCT/US2003/013917 WO2004062569A2 (fr) 2003-01-07 2003-05-04 Compositions de cardiolipine, leurs procedes de preparation et d'utilisation
WOPCT/US03/16412 2003-05-23
PCT/US2003/016412 WO2003099830A2 (fr) 2002-05-24 2003-05-23 Compositions de cardiolipine, leurs procedes de preparation et d'utilisation
PCT/US2003/027806 WO2004039817A1 (fr) 2002-10-16 2003-09-05 Molecules de cardiolipine et procedes pour leur synthese
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US8815599B2 (en) 2004-06-01 2014-08-26 Pronai Therapeutics, Inc. Methods and compositions for the inhibition of gene expression
US20080286351A1 (en) * 2004-06-29 2008-11-20 Ahmad Moghis U Pegylated Cardiolipin Analogs, Methods of Synthesis, and Uses Thereof
CA2587103A1 (fr) * 2004-11-08 2006-05-18 Neopharm, Inc. Synthese de produits analogues de la cardiolipine et leurs applications
EP1874793A4 (fr) 2005-04-15 2008-12-24 Univ Texas Administration d'arnsi par compositions lipidiques neutres
WO2007014150A2 (fr) * 2005-07-22 2007-02-01 Neopharm, Inc. Procede d'administration de liposomes contenant des oligonucleotides
ES2419106T3 (es) 2005-12-01 2013-08-19 Pronai Therapeutics, Inc. Formulación de liposomas anfóteros
WO2007065017A2 (fr) * 2005-12-01 2007-06-07 Pronai Therapeutics, Inc. Systeme d'administration de liposomes cationiques a oligonucleotides
MX2008010826A (es) * 2006-02-24 2009-03-02 Neopharm Inc Metodo y proceso para preparar cardiolipina.
US9750812B2 (en) * 2008-09-27 2017-09-05 Jina Pharmaceuticals, Inc. Lipid based pharmaceutical preparations for oral and topical application; their compositions, methods, and uses thereof
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