WO1994008563A1 - Classe nouvelle de derives de phosphocholine a activite fongicide - Google Patents

Classe nouvelle de derives de phosphocholine a activite fongicide Download PDF

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Publication number
WO1994008563A1
WO1994008563A1 PCT/US1993/009623 US9309623W WO9408563A1 WO 1994008563 A1 WO1994008563 A1 WO 1994008563A1 US 9309623 W US9309623 W US 9309623W WO 9408563 A1 WO9408563 A1 WO 9408563A1
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alkyl
alkynyl
alkenyl
branched
radicals
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PCT/US1993/009623
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English (en)
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WO1994008563A9 (fr
Inventor
Michael Tempesta
Shivanand D. Jolad
Steven King
Guohua Mao
Reimar C. Bruening
John E. Kuo
Thien V. Truong
Donald E. Bierer
Jeffrey M. Dener
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Shaman Pharmaceutical, Inc.
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Priority to US08/411,709 priority Critical patent/US5681829A/en
Priority to AU53544/94A priority patent/AU5354494A/en
Priority to JP6510144A priority patent/JPH08502298A/ja
Priority to EP93923809A priority patent/EP0663816A4/fr
Publication of WO1994008563A1 publication Critical patent/WO1994008563A1/fr
Publication of WO1994008563A9 publication Critical patent/WO1994008563A9/fr

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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
    • C07F9/093—Polyol derivatives esterified at least twice by phosphoric acid groups
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N57/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds
    • A01N57/10—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds having phosphorus-to-oxygen bonds or phosphorus-to-sulfur bonds
    • A01N57/14—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds having phosphorus-to-oxygen bonds or phosphorus-to-sulfur bonds containing aromatic radicals
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N57/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds
    • A01N57/10—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds having phosphorus-to-oxygen bonds or phosphorus-to-sulfur bonds
    • A01N57/16—Biocides, pest repellants or attractants, or plant growth regulators containing organic phosphorus compounds having phosphorus-to-oxygen bonds or phosphorus-to-sulfur bonds containing heterocyclic radicals
    • 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
    • A61K31/66—Phosphorus compounds
    • A61K31/683—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols
    • A61K31/685—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols one of the hydroxy compounds having nitrogen atoms, e.g. phosphatidylserine, 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
    • A61K31/66—Phosphorus compounds
    • A61K31/683—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols
    • A61K31/688—Diesters of a phosphorus acid with two hydroxy compounds, e.g. phosphatidylinositols both hydroxy compounds having nitrogen atoms, e.g. sphingomyelins
    • 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
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • 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
    • A61P31/10—Antimycotics
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
    • C07H15/02—Acyclic radicals, not substituted by cyclic structures
    • C07H15/04—Acyclic radicals, not substituted by cyclic structures attached to an oxygen atom of the saccharide radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
    • C07H15/18—Acyclic radicals, substituted by carbocyclic rings

Definitions

  • This invention relates to new classes of phosphocholine derivatives as well as to various methods for preparing these compounds — including synthetic, enzymatic and extractive using certain plants.
  • the phosphocholine derivatives of the invention are non-toxic and exhibit substantial antifungal activity in slowing fungal growth and in killing fungi.
  • the plant species Irlbachia alata has been used as an anti-infective agent in the Peruvian Amazon region.
  • the leaves are squeezed and the liquid is applied to infected skin sores.
  • the same liquid from the leaves is applied to skin problems and skin fungal infections. It is utilized to treat vaginal yeast infections.
  • Irlbachia alata is one species of 10-12 species of the plant family Gentianaceae . These species occur in tropical South America especially in the Amazon and Negro River basins. The plants in the genus Irlbachia are generally low herbs characteristically with 3-5 plinerved leaves. The most consistent diagnostic feature for the genus is the pollen morphology. A reference to Irlbachia alata and related species was made in 1775 by the French scientist Fusee Aublet (Aublet, F. 1775, aria d.es Plantes de la Guiane Francoise, Didot, Paris) . The ethnobotanical notes from this reference were subsequently compiled and republished in English. Aublet noted the following about two species in the genus Irlbachia :
  • these compounds are phosphocholine derivatives (1 or 2-deacyl-phosphatidyl cholines) in which the 1 or 2-OH-group of the glycerol moiety has been glycosylated with glucose, galactose, arabinose, mannose, rhamnose or another sugar.
  • the basic chemical structure may be drawn as follows:
  • R or R 1 is a sugar moiety and the other is an acyl or sugar moiety.
  • the molecular backbone common to all members of this class of compounds is drawn above.
  • the acyl- group can be any long-chain fatty acid, while the sugar unit can be any of the sugars commonly found in plants, including but not limited to glucose, galactose, arabinose, mannose, rhamnose, or another naturally occurring sugar.
  • Q is C2 to C30 alkyl, alkenyl, alkynyl, branched alkyl, branched alkenyl, or branched alkynyl; _ ⁇ _
  • Z is oxygen or sulfur;
  • X and Y are independent oxygen, sulfur, CH 2 , CF 2 , or N-R,;
  • A, B, and T are independently alkyl, alkenyl, alkynyl, branched alkyl, branched alkenyl, or branched alkynyl radicals of Cl to C20 chain lengths; are independently or together cycloalkyl or bridged cycloalkyl radicals of ring size C3 to C20, or cylcoalkenyl, bridged cycloalkenyl or cyclo(polyene)radicals of ring size C4 to C20, cycloalkynyl, bridged cycloalkeynl or cyclo(polyalkynyl)radicals of ring size C8 to C20;
  • D is oxygen, sulfur, CH 2 , CF 2 , or N-R 2 ;
  • F is alkyl, alkenyl, alkynyl, branched alkyl, branched alkenyl, branched alkynyl, cycloalkyl, bridged cycloalkyl, cycloalkenyl or cycloalkynyl radicals containing Cl to C20 carbon atoms;
  • R, and R 2 are independently hydrogen, alkyl, alkenyl, alkynyl, branched alkyl, branched alkenyl, branched alkynyl, cycloalkyl, bridged cycloalkyl, cycloalkenyl, bridged cycloalkenyl or cycloalkynyl radicals containing Cl to C20 carbon atoms, or any protecting group described in the book "Protecting Groups in Organic Synthesis" by Theodora Greene and Peter G.M. Wuts.
  • the central carbon atom can be either the R and S optical stereoisomer or a mixture of R and S stereoisomers, and where AA, BB, and CC are defined as follows: where AA, is A-J with A being attached to the carbon atom of the three carbon central unit and J is defined below;
  • BB is B-Y, with B being attached to the carbon atom of the three carbon central unit and Y is defined below:
  • A is oxygen, sulfur, CH 2 , CF 2 or N-R
  • B is oxygen, sulfur, CH 2 , CF 2 or N-R 2
  • D is oxygen, sulfur, CH 2 , CF 2 or N-R 3
  • Y is alkyl, alkenyl, alkynyl, poly(alkenyl) , poly(alkynyl) , or poly(alkenoalkynyl) radicals comprised of Cl to C20 carbon atoms chain lengths, or alkanoyl, alkenoyl, _alkynoyl, poly(alken)oyl, poly(alkyn)oyl or poly(alkenoalkyn)oyl radicals comprised of C2 to C20 chain lengths or alkyloxy, alkenyloxy, alkynyloxy, poly(alkenyl)oxy, poly(alkynyl)oxy, poly(alkenoalkynyl)oxy radicals comprised of Cl to C20 carbon atoms
  • X is oxygen, sulfur, CH 2 , CF 2 or N-R 4 ;
  • F, K, L and M are independently hydrogen, hydroxyl, protected hydroxyl (as described in the book “Protecting Groups in Organic ' Synthesis” by Theodora Greene and Peter G.M.
  • Z is oxygen or sulfur
  • E is oxygen, sulfur, CH 2 CF 2 or N-R s ;
  • G is alkyl, branched alkyl, cycloalkyl or bridged cycloalkyl radicals of Cl to C20 chain lengths;
  • Q is halogen, hydroxyl, protected hydroxyl utilizing any protecting groups described in the book “Protecting Groups in Organic Synthesis” by Theodora Greene and Peter G.M.
  • Ri, R 2 , R 3/ R 4 and Rj are independently alkyl, alkenyl, alkynyl, branched alkyl, branched alkenyl, branched alkynyl, cycloalkyl, bridged cycloalkyl, cycloalkenyl or cycloalkynyl radicals of Cl to C20 chain lengths, or any protecting group described in the book "Protecting Groups in Organic Synthesis" by Theodora Greene and Peter G «M.
  • Rj is phenyl or phenylmethyl, hydrogen, or nil
  • R 2 is hydrogen, phenylmethyl, or any protecting group described in the book “Protecting Group in Organic Synthesis” by Theodora Green and Peter G.M. Wuts which can be cleaved by hydrogenolysis;
  • AA, BB, and Q are as defined above where the central carbon atom of the three carbon unit is either the R optical isomer, the S optical isomer, or any mixture of the two optical isomers thereof;
  • Another preferred subgroup of the above-described Class III of phosphocholine derivatives have the following structures:
  • R is phenyl or phenylmethyl, hydrogen, or nil
  • R 2 is hydrogen, phenyl methyl or any protecting group described in the book “Protecting Groups in Organic Synthesis” by Theodora Greene and Peter G.M. Wuts which can be cleaved by hydroge olysis;
  • R 3 is hydrogen or a protecting group as described in the book “Protecting Groups in Organic Synthesis” by Theodora Greene and Peter G.M. Wuts.; where the central carbon atom of the three carbon unit is either the R optical isomer, the S optical isomer, or any mixture of the two optical isomers thereof; and Q is defined above.
  • R is phenyl or phenylmethyl, hydrogen, or nil
  • R 2 is a protecting group as described in the book “Protecting Groups in Organic Synthesis” by Theodora Greene and Peter G.M. Wuts, or hydrogen if R ! is not hydrogen; and Q is defined above.
  • Fig. 1 is the FTIR spectrum of the composition comprising a phosphocholine derivative obtained from Irlbachia alata.
  • Fig. 2 is the proton NMR spectrum of the composition comprising a phosphocholine derivative obtained from JrlJac ia alata in D 2 0 at 400 mHz.
  • Fig. 3 is the FAB"/MB mass spectrum of the composition comprising a phosphocholine derivative obtained from Irlbachia alata .
  • the glysosylated lysolecithins of the invention can be prepared by synthetic methods or by enzymatic methods.
  • the phosphocholine derivatives can be prepared either by synthetic methods or by methods entailing extraction from plant materials.
  • benzyl ethers or the benzilidine moiety are the preferred protecting group, since they can be selectively removed by catalytic hydrogenation, while leaving the sensitive acyl-glycerol linkage intact.
  • the glycosidation requires silver, mercury (Helferich modification) , or cadmium salts as catalytic halogen abstractor, in the presence of a dehydrating agent (Timell, T.E., Can .J. Chem . 1964, 4_2., 1456; Dejter-Juszynsky, M. and Flowers, H.M. , Carbohydr. Res .
  • R ⁇ Palmitoyl-, Oleyl-, Palmitoteyl-, etc.) 1) Et ⁇ NBr/iPr ⁇ tN/Mol.Sieves; DCM/THF. 2) HJPd;EtOAc/THF.
  • glycosyltransferases As an alternative to the synthetic sequence outlined above, an in vitro enzymatic glycosidation simulating the biosynthetic process will produce the desired compounds in comparable yields.
  • the natural glycosidation catalysts are glycosyltransferases. These enzymes operate with uridinediphospho-glycosides (UDP-sugars) as substrates and ATP as the energy source. While the enzymes have to be prepared from fresh plant material, UDP-sugars, ATP, as well as the respective phosphocholine derivatives are commercially available. This synthesis has the advantage of being essentially a one-step process with the high selectivity and yields expected from an enzymatic reaction. The following scheme describes the preparation of a glucoside. Other transferases, not specific to glucose, could be applied in the preparation of glycosylated lysolecithins with other sugars as well:
  • An alcohol is phosphorylated or glycosylated.
  • the product is subsequently deprotected.
  • the deprotected product is then alkylated or esterified to produce the phosphocholine derivatives.
  • the general scheme for this outlined synthetic method is shown below.
  • Glycerol Derivative 0 can be either the R or S optical isomer, racemic, or a mixture of R and S is ⁇ mers i implies that a number of synthetic transformations are required
  • R-i Sugar, carbocyclic sugar, functionalized sugar derivative, etc.
  • R 3 alkyl, alkanoyl, alkenyl, alkenoyl, etc.
  • X, Y, and Z can be C, O, N, S independently or equal to each other
  • the phosphocholine derivative in Classes I, II and III are all useful in treating fungal infection by the administration to a warm-blooded animal of a therapeutically effective amount of a phosphocholine derivative.
  • the pharmaceutical composition comprising the phosphocholine derivative used for such administration may also contain pharmaceutically acceptable excipients and carriers.
  • Phosphocholine derivatives in Classes I and II are believed to be novel compositions.
  • the antifungal agent of Classes I, II and III may be administered to a warm-blooded animal intravenously, intraperitoneally, subcutaneously, intramuscularly, orally, topically, by aerosol, or combinations thereof.
  • the antifungal agent of phosphocholine derivatives in Class II can be administered intravenously in a range of about 0.1 to about 10 mg/kg.
  • the fungal agent of Class II can be administered intraperitoneally in a range of about 0.1 to about 10 mg/kg.
  • the fungal agent of Class II can be administered subcutaneously in a range of about 1 to about 20.
  • the fungal agent of Class II can be administered intramuscularly in a range of about 1 to about 20.
  • the fungal agent of Class II can be administered orally in a range of about 5.0 to about 30 mg/kg.
  • the fungal agent of Class II can be administered topically in a range of about 5.0 to about 15% by weight.
  • the fungal agent of Class II can be administered by aerosol in a range of about 5.0 to about 30 mg/kg/day.
  • the above dosage ranges may need to be doubled for those phosphocholine derivatives in Class I and III with lower antifungal activity which are identical or similar to those in table 2 (see below) .
  • Plants are not known to contain phosphocholine derivatives.
  • the plant source material such as the whole plant, the roots, leaves, stem and/or latex of the plant, is extracted with water and/or a water miscible solvent.
  • the preferred solvents are alcohol of 1-3 carbon atoms or acetone.
  • the aqueous extract is extracted with butanol.
  • the butanol-soluble fraction is subjected to gel filtration (e.g., over Sephadex), reversed-phase column chromatography (e.g., C-8) , or gel-permeation chromatography (e.g., divinyl benzene cross-linked gels) such as PL-GEL or membranes (e.g., an Amicon membrane) using water or water and a water miscible solvent, with or without a buffer, as the mobile phase.
  • the water miscible solvent is preferably a 1-3 carbon alcohol, acetone or acetonitrile.
  • the useful phosphocholine derivatives containing compound is the fraction detected by NMR spectroscopy.
  • a specific member of the class of phosphocholine derivatives of the present invention is - 20 -
  • 2-palmitoyl-l-0- glucopyranosyllysolecithin is a relatively active antifungal agent similar in activity to L-a-Lysophosphatidyl inositol, discussed in Table 2 below.
  • the 5 isopropanol/water (1:1 v/v) soluble extract was partitioned between water and ethyl acetate.
  • the ethyl acetate phase was separated and discarded.
  • the water soluble phase after extraction with n-butanol, was then discarded.
  • the n-butanol phase was subjected o to filtration over two Sephadex LH-20 gel columns using 90% aqueous ethanol (for first filtration) and 20% aqueous acetone (for second filtration) as the mobile phases.
  • 1,22-docosandiol bisphosphocholine ester was collected from the early fractions of each 5 gel filtration.
  • Chelonanthus alatus There are several uses described for Chelonanthus alatus , including oral decoctions to treat smallpox, fevers and for gastric disturbances.
  • the isolated phosphocholine derivative fraction containing 1,22-docosandiol bisphosphocholine ester has the characteristic IR, proton NMR and FAB" mass spectra shown in Figs. 1, 2 and 3, respectively.
  • the IR spectrum has peaks at approximately 1060, 1220, 1475, 1600-1700, 2850, 2950 and 3400 cm '1 .
  • the ! H NMR spectrum has major peaks at ⁇ 1.2, 1.4, 1.7, 3.1, 3.5, 3.7 and 4.3.
  • the FAB"/MB mass spectrum has major peaks (>40%) at m/z 657, 612, 587, 586, 555, 493, 491, 475, 403, 277, 233, 201, 194, 179, 168, 165 and 163.
  • the high resolution mass spectrum (FAB + ) has a molecular ion at 673.4669 amu.
  • Tetrahydrofuran was distilled from potassium/benzophenone; benzene, triethylamine, and methylene chloride, N-methylmorpholine, and benzyl alcohol were distilled from calcium hydride; 2- bromoethylphosphorodichloridate was prepared according to the procedure reported by Bau ann et al Lipids, 17, 453 (1982) and was freshly distilled prior to use; trifluromethanesulfonic anhydride was freshly distilled under inert atmosphere; O- ⁇ -D- (Glucopyranosyl)trichloroacetimidate was prepared by the method of Schmidt.
  • Trifluoromethanesulfonic anhydride (1.25 mL, 7.6 mmol) was then added, and the reaction was stirred for 30 minutes with the temperature maintained at 5°C. The solution was then filtered through a bed of silica. The filtrate was concentrated under reduced pressure at 30°C to give an orange/brown oil (1.84 g, 7.0 mmol) in 92% yield which was used directly for the next step.
  • 2,3,4,6-Tetra-O-benzyl-D-glucopyranose 100 g, 0.182 mol was dissolved in THF (1.4 L) and chilled to -10°C in a nitrogen-purged 3-L three-necked morton flask fitted with a thermometer, stopper, and mechanical stirrer.
  • Sodium hydride 60% in oil (16.1 g, -0.403 mol) was added in 4 increments over 10 minutes, and the solution was stirred for 30 minutes.
  • the reaction mixture was stirred for 11 h at O o c, left to sit at -15°C for 12 h, and stirred again for 4 h at rt.
  • Water (100 mL) was added, and the solution was concentrated to 200 L of solution.
  • the concentrate was redissolved in methylene chloride (750 mL) in a 3-L separatory funnel and washed with water three times (750 mL, 2 x 500 mL) .
  • the combined aqueous layers were extracted with diethyl ether (500 mL) .
  • the combined organic layers were concentrated to give a red oil which was purified by flash chromatography (silica gel, 33-40% gradient of ethyl acetate/hexane).
  • Procedure B Compound 4 (500 mg, 0.52 mmol) was dissolved in THF (20 mL) in a 100-mL three-necked round-bottomed flask fitted with two stoppers and a septum. Glacial acetic acid (9.5 mL) was added, and the solution was chilled to 0°C. A solution of TBAF (5.16 mL, 1.0 M in THF) was syringed into the chilled solution, and stinting was continued at 0°C for 8 h and—then at rt for 25 hours.
  • the solution was concentrated under reduced pressure to approximately 30 mL of liquid and then redissolved in methylene chloride (150 mL) .
  • the organic layer was washed with water (3 x 120 inL) a ⁇ d neutralized with sodium bicarbonate solution (2 x 150 mL) .
  • the combined aqueous layers were extracted with methylene chloride (100 mL) .
  • the combined organic layers were dried over magnesium sulfate, filtered, and concentrated.
  • the resulting dark red concentrate was purified by flash chromatography (silica gel, 25% ethyl acetate/hexane) to _give 6 a colorless oil which corresponded to an upper TLC spot (1.3 g, 1.52 mmol) in 46% yield.
  • Procedure A In a nitrogen-purged 100-mL three-necked round-bottomed flask fitted with two stoppers and a o septum was dissolved freshly distilled 2- bromoethylphosophorodichloridate (1.72 g, 7.11 mmol) in diethyl ether (20 mL) . The solution was chilled to 0°C, and triethylamine (8.15 mL, 58.5 mmol) was injected into the solution which caused precipitation 5 of a white solid. A solution of compound 5 (1.0 g,
  • Procedure B In a nitrogen-purged 100-mL three-necked roundbottomed flask fitted with a thermometer, stopper, and septum was dissolved freshly distilled 2- bromoethylphosphorodichloridate (1.42 g, 5.85 mmol) in methylene chloride (15 mL) . The solution was chilled to 0°C, and compound 5 (1.0 g, 1.17 mmol) and a solution of N-methylmorphiline (1.28 mL, 11.7 mmol) dissolved in methylene chloride (35 mL) was injected into the solution over a 10 minute period.
  • reaction mixture was stirred at 0°C for 5.5 h at which point a new TLC spot which co-spotted with secondary alcohol 6 appeared. Stirring was continued for another 30 minutes, and benzyl alcohol (1.21 ml, 11.7 mmol) was injected into the reaction. After 6 days of stirring, the reaction mixture was transferred to a 500-mL separatory funnel, and methylene chloride (150 mL) and water (200 ml) were added. The layers were separated, and the organic layer was rotary evaporated under reduced pressure.
  • reaction mixture was stirred under nitrogen overnight, transferred to a 1-L separatory funnel, and chloroform (300 mL) and water (300 mL) were added. The aqueous layer was extracted with chloroform (2 x 100 mL) and then the combined organic layers were washed with water (3 x 100 mL) .
  • Glacial acetic acid (13.8 mL 241 mmol) was then poured into the reaction mixture to quench the reaction, and the resulting solution was stirred for approximately 30 minutes.
  • the reaction mixture was poured into a separatory funnel containing ice water (500 mL) and methylene chloride (200 mL) . The layers were separated, and aqueous layer was extracted twice more with methylene chloride (100 mL portions) and then the combined organic layer was washed with brine (400 mL) .
  • 2-brorhoethylphosphorodichloridate (17.2 g, 71.1 mmol) in anhyd diethyl ether (500 mL) .
  • the solution was chilled to 0°C and triethylamine (81.5 mL, 0.585 mol) was injected into the solution, causing precipitation of a white solid.
  • a solution of 12 (10.0 g, 11.7 mmol) dissolved in diethyl ether (250 mL) was cannulated into the morton flask, and the solution was stirred for 1.5 hours. TLC showed disappearance of 12.
  • Benzyl alcohol (12.1 mL, 0.117 mol) was injected into the reaction mixture, and stirring was continued at rt for 16 h.
  • reaction mixture was rotary evaporated to a small volume and purified by flash chromatography (silica gel, diethyl ether:hexane, 1:3 to elute UV-nonactive impurities, diethyl ether:hexane,1:1 to elute compound 22).
  • reaction mixture was evaporated to a small volume and separated by flash chromatography (silica gel, diethyl ether/hexane, 1:3) to give compound 17 as a colorless oil (120 mg, 22%) , which was identical to the material described earlier.
  • the antifungal activity of the isolated phosphocholine fraction was determined in vitro by using three fungal cultures — Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus .
  • the method used to determine in vitro antifungal activity is discussed in McGinnis, M.R., Laboratory Handbook of Medical Mycology, Academic Press, New York, London, p66l (1980); and Droughet E., Dupont, B.7 ⁇ Improvisi, L. , Vivian, M.A.
  • R is the group identified in table 2.
  • the toxicity of the isolated phosphocholine derivative fraction is low, based on tests with Hep 2 cells indicating an ID 50 of greater than 1000 ug/ml.
  • the method used in determining cytotoxicity is discussed in Mosmann, T. , "Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays", J. Immun . Methods , 65, 55-63, 1986.
  • the isolated fraction having the above-described in vitro antifungal activity and low toxicity is expected to similarly exhibit significant in vivo antifungal activity against fungal infections which are dermatophytic, systemic, ophthalmic and vaginal.
  • Other human and animal infections treatable with the compounds of the present invention include aspergilliosis, candidiasis, and cryptococcus infections.

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Abstract

Certains dérivés de phosphocholine ont une activité fongicide thérapeutique considérable. Ces dérivés de phosphocholine peuvent être synthétisés chimiquement, préparés par voie enzymatique ou extraits de la plante Irlbachia alata. Ces dérivés de phosphocholine sont utiles pour traiter des infections par des champignons, y compris des infections dermatophytiques, systémiques, ophtalmiques et vaginales.
PCT/US1993/009623 1992-10-08 1993-10-08 Classe nouvelle de derives de phosphocholine a activite fongicide WO1994008563A1 (fr)

Priority Applications (4)

Application Number Priority Date Filing Date Title
US08/411,709 US5681829A (en) 1992-10-08 1993-10-08 Class of phosphocholine derivatives having antifungal activity
AU53544/94A AU5354494A (en) 1992-10-08 1993-10-08 Novel class of phosphocholine derivatives having antifungal activity
JP6510144A JPH08502298A (ja) 1992-10-08 1993-10-08 抗真菌活性をもつ新規クラスのホスホコリン誘導体
EP93923809A EP0663816A4 (fr) 1992-10-08 1993-10-08 Classe nouvelle de derives de phosphocholine a activite fongicide.

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US95841692A 1992-10-08 1992-10-08
US958,416 1992-10-08

Publications (2)

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WO1994008563A1 true WO1994008563A1 (fr) 1994-04-28
WO1994008563A9 WO1994008563A9 (fr) 1994-06-09

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PCT/US1993/009623 WO1994008563A1 (fr) 1992-10-08 1993-10-08 Classe nouvelle de derives de phosphocholine a activite fongicide

Country Status (5)

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EP (1) EP0663816A4 (fr)
JP (1) JPH08502298A (fr)
AU (1) AU5354494A (fr)
CA (1) CA2146639A1 (fr)
WO (1) WO1994008563A1 (fr)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0796112A1 (fr) * 1994-11-30 1997-09-24 Amur Research Corp. Derives medicamenteux de la phosphocholine
US5681829A (en) * 1992-10-08 1997-10-28 Shaman Pharmaceuticals, Inc. Class of phosphocholine derivatives having antifungal activity
US5811568A (en) * 1992-10-08 1998-09-22 Shaman Pharmaceuticals, Inc. Process for the preparation of mono- and bis(phosphocholine) derivatives which have antifungal activity
EP1389909A1 (fr) * 2001-05-03 2004-02-25 Oligos Etc. Inc. Composes antimicrobiens et leurs procedes de mise en oeuvre
US7868162B2 (en) 1998-12-30 2011-01-11 Lakewood-Amedex, Inc. Antimicrobial and antiviral compounds and methods for their use
US8435960B2 (en) 1998-12-30 2013-05-07 Lakewood-Amedex, Inc. Devices for improved wound management

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4684625A (en) * 1982-07-08 1987-08-04 Syntex (U.S.A.) Inc. Method for enhancing the anti-infective activity of muramyldipeptide derivatives

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE3829899C2 (de) * 1988-09-02 1994-12-15 Reutter Werner Glyceringlycophosphatide und diese enthaltende pharmazeutische Mittel zur Bekämpfung von Hauterkrankungen

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4684625A (en) * 1982-07-08 1987-08-04 Syntex (U.S.A.) Inc. Method for enhancing the anti-infective activity of muramyldipeptide derivatives

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
See also references of EP0663816A4 *
Tetrahedron Letters, Volume 37, issued 1979, VAN BOECKEL et al., "Synthesis of Glucosyl Phosphatidylglycerol via a Phosphotriester Intermediate", pages 3561-3564, especially page 3563. *

Cited By (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5681829A (en) * 1992-10-08 1997-10-28 Shaman Pharmaceuticals, Inc. Class of phosphocholine derivatives having antifungal activity
US5811568A (en) * 1992-10-08 1998-09-22 Shaman Pharmaceuticals, Inc. Process for the preparation of mono- and bis(phosphocholine) derivatives which have antifungal activity
EP0796112A1 (fr) * 1994-11-30 1997-09-24 Amur Research Corp. Derives medicamenteux de la phosphocholine
EP0796112A4 (fr) * 1994-11-30 1999-12-01 Amur Research Corp Derives medicamenteux de la phosphocholine
US7176191B2 (en) 1998-12-30 2007-02-13 Oligos Etc. Inc. Antimicrobial compounds and methods for their use
US7868162B2 (en) 1998-12-30 2011-01-11 Lakewood-Amedex, Inc. Antimicrobial and antiviral compounds and methods for their use
US8435960B2 (en) 1998-12-30 2013-05-07 Lakewood-Amedex, Inc. Devices for improved wound management
EP1389909A1 (fr) * 2001-05-03 2004-02-25 Oligos Etc. Inc. Composes antimicrobiens et leurs procedes de mise en oeuvre
EP1389909A4 (fr) * 2001-05-03 2005-03-30 Oligos Etc Inc Composes antimicrobiens et leurs procedes de mise en oeuvre
AU2002305336B2 (en) * 2001-05-03 2007-11-08 Lakewood-Amedex, Inc. Antimicrobial compounds and methods for their use
EP2311466A3 (fr) * 2001-05-03 2012-04-25 Lakewood-Amedex, Inc Composés antimicrobiens et leurs procédés de mise en oeuvre

Also Published As

Publication number Publication date
CA2146639A1 (fr) 1994-04-28
AU5354494A (en) 1994-05-09
JPH08502298A (ja) 1996-03-12
EP0663816A4 (fr) 1996-04-03
EP0663816A1 (fr) 1995-07-26

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