WO1999039201A1 - GLYCOSYLATION DE SULFOXYDE EN SOLUTION ET EN PHASE SOLIDE: SYNTHESE D'OLIGOSACCHARIDES A LIAISON β A L'AIDE DE DONNEURS 2-DEOXY-2-N-TRIFLUOROACETAMIDO-GLYCOPYRANOSYLE - Google Patents

GLYCOSYLATION DE SULFOXYDE EN SOLUTION ET EN PHASE SOLIDE: SYNTHESE D'OLIGOSACCHARIDES A LIAISON β A L'AIDE DE DONNEURS 2-DEOXY-2-N-TRIFLUOROACETAMIDO-GLYCOPYRANOSYLE Download PDF

Info

Publication number
WO1999039201A1
WO1999039201A1 PCT/US1999/002180 US9902180W WO9939201A1 WO 1999039201 A1 WO1999039201 A1 WO 1999039201A1 US 9902180 W US9902180 W US 9902180W WO 9939201 A1 WO9939201 A1 WO 9939201A1
Authority
WO
WIPO (PCT)
Prior art keywords
deoxy
trifluoroacetamido
glycosyl
phenylsulfenyl
acetyl
Prior art date
Application number
PCT/US1999/002180
Other languages
English (en)
Inventor
Domingos Silva
Michael J. Sofia
Original Assignee
Advance Medicine Est, Inc.
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Advance Medicine Est, Inc. filed Critical Advance Medicine Est, Inc.
Priority to EP99905611A priority Critical patent/EP1053471A1/fr
Priority to CA002319339A priority patent/CA2319339A1/fr
Priority to AU25736/99A priority patent/AU2573699A/en
Priority to JP2000529604A priority patent/JP2002501932A/ja
Publication of WO1999039201A1 publication Critical patent/WO1999039201A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H5/00—Compounds containing saccharide radicals in which the hetero bonds to oxygen have been replaced by the same number of hetero bonds to halogen, nitrogen, sulfur, selenium, or tellurium
    • C07H5/08—Compounds containing saccharide radicals in which the hetero bonds to oxygen have been replaced by the same number of hetero bonds to halogen, nitrogen, sulfur, selenium, or tellurium to sulfur, selenium or tellurium
    • C07H5/10—Compounds containing saccharide radicals in which the hetero bonds to oxygen have been replaced by the same number of hetero bonds to halogen, nitrogen, sulfur, selenium, or tellurium to sulfur, selenium or tellurium to sulfur
    • 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
    • C07H15/12—Acyclic radicals, not substituted by cyclic structures attached to a nitrogen atom of the saccharide radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B2200/00—Indexing scheme relating to specific properties of organic compounds
    • C07B2200/11—Compounds covalently bound to a solid support
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B40/00—Libraries per se, e.g. arrays, mixtures

Definitions

  • the present invention generally relates to ⁇ -oligosaccharides and a process for the synthesis of ⁇ -oligosaccharides comprising reacting a glycosyl donor and a glycosyl acceptor. More particularly, the present invention relates to a process for the synthesis of ⁇ - oligosaccharides using 2-deoxy-2-N-trifluoroacetamido glycopyranosyl sulfoxide as a glycosyl donor, in solution and solid phase sulfoxide glycosylations. The present invention also relates to design, construction and analysis of a disaccharide combinatorial library.
  • glycosidic residues act as recognition signals that mediate key events in normal cellular development and function. They are involved in embryogenesis, hormonal activities, neuronal development, inflammation, cellular proliferation, fertilization and the organization of different cell types into specific tissues. They are able to regulate the transportation of proteins between cells and should be regarded as signal substances in metabolism.
  • Oligosaccharides are also involved in the prevention and treatment of diseases.
  • Compounds of this class include glycosamine and macrolide antibiotics, anthracycline and enediyne anticancer antibiotics.
  • oligosaccharides on cell surfaces function as receptors for viruses, toxins, infectious bacteria, hormones, pathogens, enzymes, proteins, as well as more benign ligands.
  • Oligosaccharide synthesis is by far one of the most challenging fields in modern organic chemistry. Efficient construction of an oligosaccharide or a glycoconjugate involves attaching a glycosyl moiety to a specific position of a glycosyl acceptor not only in high yield but also with high stereocontrol. Based on this challenge, several glycosylation methods have been developed. 3
  • Thioglycosides have attracted considerable attention as glycosyl donors. Thioalkyl or aryl groups offer efficient temporary protection of the anomeric center and, at the same time, can be selectively activated under various conditions. Many glycosylation reactions using thioglycosides have been reported. Early attempts to use thioglycosides as glycosyl donors include activation by mercury (II) sulfate, Ferrier et al., J Glycoconjugate, 4:97-108 (1987); mercury (II) chloride, Tsai et al., Can.
  • the first is a "two step" activation which involves first forming a glycosyl halide, and then further activating this with a halophilic reagent. Sato et al, Carbohyd. Res., 155:C6-10 (1986).
  • Soc, 106:4189 (1984) employs a stable phenylthioglycoside as the key building block.
  • the phenylthioglycoside is converted to the more reactive glycosyl fluoride by treatment with NBS and diethyl amino sulfur trifluoride (DAST).
  • DAST diethyl amino sulfur trifluoride
  • the glycosyl fluoride is coupled with the glycosyl acceptor which carries phenylthio at the anomeric position in the preparation of oligosaccharide chains.
  • the two stage activation procedure is particularly suited for solid phase oligosaccharide synthesis.
  • the second technique involves a one step activation with a thiophilic reagent such as methyl triflate or dimethyl(methylthio)sulfonium trifluoromethane sulfonate (DMTST).
  • a thiophilic reagent such as methyl triflate or dimethyl(methylthio)sulfonium trifluoromethane sulfonate (DMTST).
  • DMTST is a highly thiophilic promoter in the synthesis of 1,2 trans glycosides using various thioglycosides with participating 2-substituents as glycosyl donors. Fugedi et al., J. Glycoconjugate, 4:97-108 (1987).
  • Another method of thioglycoside activation developed by Sinay et al., Pure Appl. Chem. 63:519 (1991) involves electron transfer from sulfur to the activating agent tris( ⁇ - bromophenyl) ammoniumyl hexachloroantimonate (TBPA).
  • TBPA tris( ⁇ - bromophenyl) ammoniumyl hexachloroantimonate
  • the generated glycosyl radical cation suffers cleavage to a thiyl radical, leaving behind an oxonium species which then undergoes glycosidation.
  • the sulfoxide glycosylation method of the present invention has been shown to be successful in both solution and solid phases. It allows the glycosylation of sensitive and/or unreactive substrates at low temperatures and under essentially neutral conditions, with high degree of stereoselectivity in general.
  • the present invention is generally directed to a process for the synthesis of ⁇ - oligosaccharides, including ⁇ -disaccharides and their conjugates, which process comprises reacting a glycosyl donor and a glycosyl acceptor.
  • the invention is directed to a process for the synthesis of ⁇ -oligosaccharides using alkylsulfenyl- or arylsulfenyl- 2-deoxy-2-N-trifluoroacetamido glycopyranose as a glycosyl donor in a sulfoxide-mediated glycosylation reaction whether in solution or in the solid phase.
  • Glycosyl donors including 7
  • a further aspect of the invention is to provide a process for constructing glycosidic linkages using arylsulfenyl- or alkylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranose as glycosyl donors.
  • ⁇ -anomers i.e., ⁇ -glycosidic linkages
  • ⁇ -glycosidic linkages can be produced on a solid phase using anomeric sugar sulfoxides as glycosyl donors.
  • the process of the present invention may also be applied to the preparation of specific oligosaccharides or glycoconjugates or to the preparation of mixtures of various oligosaccharides or glycoconjugates for the creation of glycosidic chemical libraries that can subsequently be screened to detect compounds having a desired biological activity.
  • the invention is directed to a process for the synthesis of ⁇ -glycosides in high yield in the substantial absence of ⁇ -isomers.
  • the invention relates broadly to ⁇ -selectivity in glycosylation using a glycosyl donor with a C-2 protecting group capable of neighboring group participation, such as an amide, an ester, an imide or a carbamate.
  • a glycosyl donor with a C-2 protecting group capable of neighboring group participation such as an amide, an ester, an imide or a carbamate.
  • Another aspect of the invention relates to the design, construction and analysis of a combinatorial library comprising a plurality of the novel compounds of the invention, including the salts and conjugates thereof, preferably one bound to a solid phase.
  • Figure 1 illustrates a process for the synthesis of phenylsulfenyl-2-deoxy-2-N- trifluoroacetamido-3,4,6-tri-O-acetyl- ⁇ -D-glucopyranose (6).
  • Figure 2 illustrates a process for the synthesis of phenylsulfenyl-2-deoxy-2-N- trifluoroacetamido-3,4,6-tri-O-acetyl- ⁇ -D-galactopyranose (12).
  • Figure 3 illustrates acceptors 13-15 immobilized on Rink Amide polystyrene resin.
  • Figure 4 illustrates characterized products 16-18 in the solid phase glycosylation reactions.
  • Figure 5 illustrates ⁇ NMR spectra of the reaction products which indicated that only 8
  • Figure 6 illustrates the solid-phase derivatization of 16.
  • Figure 7 illustrates analytical characterization (HPLC and LC-MS) of intermediates and final product in the derivatization of 16.
  • Figure 8 illustrates the general structure and building blocks used in the combinatorial library based on 16.
  • Figure 9 illustrates the general structure and building blocks used in the combinatorial library based on 17.
  • Figure 10 illustrates the reaction sequence used in the derivatization of 16 to generate a library.
  • Figure 11 illustrates the reaction sequence used in the derivatization of 17 to generate a library.
  • Figure 12 illustrates the LC-MS spectrum of a representative member of the combinatorial library constructed around 16.
  • Figure 13 illustrates the analytical data obtained for the library constructed around 16.
  • Figure 14 illustrates the LC-MS spectrum of a representative member of the combinatorial library constructed around 17.
  • Figure 15 illustrates the analytical data obtained for the library constructed around 17.
  • Oligosaccharide An oligomeric saccharide (or carbohydrate) containing more than one monosaccharide (or sugar) units linked through glycosidic bonds.
  • An oligosaccharide may be called a disaccharide, a trisaccharide, tetrasaccharide etc. depending on the number of monosaccharides it will yield upon hydrolysis.
  • an oligosaccharide may include its conjugates (i.e., compounds comprising the mono-, di-, tri-, etc. saccharide covalently bound to another non-sugar chemical moiety, including antitumor agents, macrolides, other natural products, amino acids, peptides, nucleosides, oligonucleotides and the like).
  • Glycoside Any sugar containing at least one pentose or hexose residue in which the anomeric carbon bears a non-hydrogen substituent.
  • the non-hydrogen substituent is a heteroatom, such as nitrogen, oxygen, phosphorus, silicon, or sulfur.
  • Glycosidic link (also glycosidic bond or linkage): The link, bond, or linkage formed by the reaction of a sugar, such as an aldose or ketose, in cyclic form with an alcohol ROH to form a mixed acetal or glycoside.
  • a sugar such as an aldose or ketose
  • ROH alcohol
  • the product is a disaccharide.
  • This disaccharide may similarly react further to form a higher oligosaccharide and eventually a polysaccharide.
  • the monosaccharide units of a polysaccharide are linked through glycoside bonds.
  • the bond may be formed by reaction of either the C-1 or C-2 hemiacetal hydroxyl with any of the hydroxyl groups of the other monosaccharide.
  • the bond may be formed in such a way that the anomeric carbon has either configuration.
  • Activating agent A chemical agent that on addition to a glycosyl sulfoxide reacts with the anomeric sulfoxide group, thus rendering the anomeric carbon susceptible to nucleophilic attack.
  • the activating agent is also able to deprotect a blocked nucleophilic group under the same conditions used to activate the anomeric sulfoxide group.
  • Glycosyl acceptor Any compound that contains at least one nucleophilic group which, under the conditions of the process of the present invention, is able to form a covalent bond with the anomeric carbon of a glycosyl donor.
  • a glycosyl acceptor is any organic molecule, including a sugar, that contains unprotected hydroxyl, amino, or mercapto groups or such groups that are blocked by protecting groups that can be removed in situ, i.e., under the reaction conditions of the present invention.
  • Glycosyl donor A sugar or glycosidic residue that bears a sulfoxide group at the anomeric carbon, which group on activation renders the anomeric carbon susceptible to attack by the nucleophilic group of a glycosyl acceptor to form the glycosidic linkage.
  • a preferred glycosyl donor is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranose. It is 10
  • the donor glycoside may be a monosaccharide, a disaccharide, a trisaccharide, etc.
  • the chemical libraries that can possibly be produced with the methods and compounds of the present invention include all forms of oligosaccharides or their conjugates or glycoconjugates.
  • Glycosidic libraries A mixture, collection, or a plurality of oligosaccharides of varying sequences which can be subjected to a screening procedure to identify compounds or molecules that exhibit biological activity.
  • Such chemical libraries may also include various conjugates or glycoconjugates.
  • Glycoconjugate Any compound or molecule that comprises a non-sugar moiety that is covalently bound to a glycosidic residue. See, also, the definition of oligosaccharide, supra.
  • Protecting group A blocking or protecting group that can be removed in situ, preferably, but not necessarily, under the same conditions used to activate an anomeric sulfoxide group. It refers to moieties ordinarily used in oligosaccharide synthesis to prevent reaction of the hydroxyl or amino groups in the reaction being conducted.
  • a preferred protecting group is the trifluoroacetyl moiety.
  • Blocking group Similar to a protecting group, a blocking group is used to prevent the inappropriate reaction of a functional group of interest.
  • the terms protecting group or blocking group can be used interchangeably.
  • substantially free may refer to the glycosylated product formed under the conditions of the present invention, which in a preferred embodiment of the invention, excludes the presence of significant amounts of ⁇ -anomer.
  • glycosyl sulfoxide donors based on galactosamine and glucosamine are investigated. More particularly, sulfoxide donors, which afford high ⁇ -selectivity, are developed. Generally, ⁇ -selectivity in glycosylation reactions is achieved by using a glycosyl donor with a C-2 protecting group capable of neighboring group participation, including but not limited to amides, esters, imidos, or carbamates.
  • this protecting group at C-2 is important not only because the sulfoxide should be reactive enough to glycosylate relatively unreactive nucleophiles, but also because this group should be easily removed to allow subsequent derivatization of the amino functionality.
  • a glycosyl acceptor is allowed to react with a glycosyl donor, which is phenylsulfenyl-2-deoxy-2-N- trifluoroacetamido-3,4,6-tri-O-acetyl- ⁇ -D-glycopyranose, either in solution or in the solid phase.
  • Another embodiment of the invention is a process for the synthesis of a glycosyl donor, which is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl- ⁇ -D- glycopyranose. These donors are found to afford the ⁇ -glycosides exclusively and in high yield.
  • the trifluoroacetamido protecting groups are then removed under mild conditions and the resulting 2-amino groups are selectively derivatized as amides.
  • this protecting group newly used in this chemistry adds flexibility to the sulfoxide glycosylation method.
  • a process for the synthesis of phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl- ⁇ -D-glycopyranose comprising the steps of: a) reacting glycosamine hydrochloride with p-methoxybenzaldehyde in the presence of alkali to form 2-N-p-methoxybenzylidene glycosamine; b) acetylating 2-N-p-methoxybenzylidene glycosamine with acetic anhydride in the presence of pyridine and dimethylaminopyridine (DMAP) to form O-acetylated 2- N-p-methoxybenzylidene glycosamine; c) removing the p-methoxy benzaldehyde group with hydrochloric acid in acetone to form O-acetylated glycosamine hydrochloride; d) protecting the
  • the preparation of the tert-butylsulfenyl counterpart can be accomplished in the same fashion by using thiotert-butanol reagent in place of thiophenol reagent .
  • the reaction solvent plays a role in the stereoselectivity of glycosylation in the absence of neighboring group participation. If a non-polar, aprotic solvent is used, the selectivity for ⁇ -glycosidic bond formation is increased while the use of a polar, aprotic solvent such as propionitrile increases selectivity for ⁇ -glycosidic bond formation.
  • the protecting groups on the glycosyl donor also have an impact on the stereochemical course of the glycosylation reaction.
  • the protecting group at the equatorial position of the C-2 center of the glycosyl donor is trifluoroacetamido, only ⁇ - glycosidic bonds are formed in the glycosylation process, regardless of whether an aprotic, non-polar solvent or an aprotic, polar solvent is used for the reaction.
  • a large number of functionalities suitable for use as protecting groups of an amino group are disclosed in T.W. Greene, Protecting Groups in Organic Synthesis, John Wiley & Sons.
  • Suitable protecting groups include carbamates such as 9-fluorenylmethoxycarbonyl (Fmoc); and allyloxycarbonyl (Alloc); imides such as phthalimido (Phth) and tetrachlorophthalimido (PhthCl 4 ); or amides such as trifluoroacetamido (TFA).
  • carbamates such as 9-fluorenylmethoxycarbonyl (Fmoc); and allyloxycarbonyl (Alloc); imides such as phthalimido (Phth) and tetrachlorophthalimido (PhthCl 4 ); or amides such as trifluoroacetamido (TFA).
  • TFA trifluoroacetamido
  • the TFA group is easily removed by treatment with LiOH in an anhydrous mixture of 50:50 MeOH-THF. This procedure is compatible with solid phase procedures. Under the conditions of the present invention, all common ester groups are also removed. This is not a drawback since amines can be selectively transformed into amides in the presence of unprotected alcohols.
  • oligosaccharides The two general methods for obtaining oligosaccharides are: a) isolation from natural sources. This approach is limited to naturally occurring oligosaccharides that are produced in 13
  • Enzymatic synthesis is limited because enzymes are highly specific and can only accept certain substrates.
  • chemical synthesis is more flexible than enzymatic synthesis and has the potential to produce an enormous variety of oligosaccharides. The problem with chemical synthesis has been that it is extremely expensive in terms of time and labor. Oligosaccharides are formed of monosaccharides connected by glycosidic linkages.
  • a fully protected glycosyl donor is activated and allowed to react with a glycosyl acceptor (typically another monosaccharide having an unprotected hydroxyl group) in solution.
  • a glycosyl acceptor typically another monosaccharide having an unprotected hydroxyl group
  • the glycosylation reaction itself can take anywhere from a few minutes to days, depending on the method used.
  • the coupled product is then purified and chemically modified to transform it into a glycosyl donor. Each purification is time consuming and can result in significant losses of material.
  • the new glycosyl donor, a disaccharide is then coupled to another glycosyl acceptor.
  • the product is then isolated and chemically modified as before.
  • solid phase synthesis of oligosaccharides requires: a) use of a saccharide derivative with a reactive leaving group at Cl; b) one hydroxyl group protected by a readily removable blocking group; c) the remaining hydroxyls protected by a stable blocking group; and d) a resin from which the formed oligosaccharide derivative can be separated without product degradation.
  • the resin has also been known to decompose due to the harshness of the conditions required for glycosylation. Furthermore, for many ester-type NPGs, there is a significant problem with acyl transfer from the glycosyl donors to the glycosyl acceptors on the resin. This side reaction caps the resin and prevents further reaction.
  • Soluble resins were employed to overcome the unfavorable reaction kinetics associated with solid-phase reactions.
  • Douglas et al., J. Am. Chem. Soc, 113: 5095 (1991) used a soluble polyethylene glycol resin with a succinic acid linker and achieved 85-95% coupling yields using the Koenigs-Knorr reaction with excellent control of anomeric stereochemistry.
  • Soluble resins may have advantages for some glycosylation reactions because they offer a more "solution-like" environment.
  • step-wise synthesis on soluble polymers requires that the intermediate be precipitated after each step and crystallized before another sugar residue can be coupled.
  • a glycosyl donor having alkyl or aryl sulfoxides at the anomeric position and a glycosyl acceptor having one or more free hydroxyls and/or other nucleophilic groups (e.g., amines) and/or silyl ether protected hydroxyls are combined in a reaction vessel.
  • glycosyl donor is blocked by a suitable protecting group such as TFA at the C-2 position resulting in a 1 , 2-trans glycosidic bond.
  • a mixture of glycosyl donors and acceptors is dissolved under anhydrous conditions in a non-nucleophilic solvent, including, but not limited to toluene, ether, tetrahydrofuran (THF), methylene chloride, chloroform, propionitrile, ethyl acetate or mixtures thereof. It has been found that the choice of solvent influences the stereochemical outcome of glycosylation for reactions in which neighboring group participation is not involved.
  • a non-nucleophilic solvent including, but not limited to toluene, ether, tetrahydrofuran (THF), methylene chloride, chloroform, propionitrile, ethyl acetate or mixtures thereof.
  • a non-polar solvent such as toluene
  • a more polar solvent such as propionitrile
  • a glycosyl acceptor is attached to an insoluble support (hereafter termed the resin) through a linkage that can be readily cleaved at the end of the synthesis using conditions that do not damage glycosidic linkages.
  • the resin may be any insoluble polymer that swells in organic solvents and has sites for attaching the glycosyl acceptor.
  • Preferred resins include, but are not limited to, polystyrene resins, such as the Merrifield resins, and PEG-derivatized polystyrene resins, such as the TentaGelTM resins.
  • the type of linkage depends on the type of functional sites available on the polymer phase and on the glycosyl acceptor.
  • the glycosyl acceptor may be any molecule having one or more reactive nucleophile including reactive hydroxyls, amines, and/or thiols, provided that it also has a suitable site for attachment to the resin.
  • a reactive nucleophile is a free nucleophile or a nucleophile with a temporal protecting group that can be removed readily once the glycosyl acceptor is attached to the resin.
  • the glycosyl acceptor may also have permanently protected nucleophiles, which are nucleophiles that cannot be deprotected under the conditions that are used to remove the temporal protecting groups.
  • the glycosyl acceptor may be a sugar or some other nucleophile-bearing molecule, including, but not limited to, steroids, amino acids or peptides, 16
  • polar lipids polycyclic aromatic compounds, macrolides, natural products and the like.
  • Preferred acceptors (13-15) immobilized on Rink amide polystyrene are shown in Figure 3.
  • the reaction mixture is diluted with 200 mL water, 200 mL aqueous NaHCO and 100 mL CH Cl .
  • the organic layer is washed with a mixture of 100 mL aqueous Na CO and 100 mL brine, dried over anhydrous Na SO and concentrated under vacuum.
  • the solid is washed with 300 mL hot ether, filtered, further washed with 300 mL of ice-cold ether and dried under vacuum, yielding 5 (48 g; 95% yield).
  • Figure 1 illustrates the synthetic scheme to arrive at the compound (6).
  • reaction mixture is left overnight at room temperature and then poured into a mixture of 25 mL methylene chloride, 15 mL saturated aqueous NaHCO , 15 mL aqueous Na CO and 15 mL brine.
  • the organic layer is further washed with a mixture of 15 mL saturated aqueous NaHCO and 15 23
  • reaction When the reaction is judged complete by TLC, it is quenched with 1 mL dimethyl sulfide and allowed to reach room temperature. The reaction mixture is diluted with 50 mL aqueous NaHCO and 50 mL CH Cl . The organic layer is
  • Figure 2 illustrates the synthetic scheme to arrive at the compound (12).
  • the glycosyl acceptor immobilized on Rink Amide resin and containing a free hydroxyl group is dried under high vacuum and then kept under argon.
  • a solution of the glycosyl donor (4 equivalents) and 2,6-di-t- butyl-4-methyl-pyridine (2 equivalents) in a solvent system compatible with the conditions of the sulfoxide glycosylation reaction generally a 9:1 mixture of anhydrous methylene chloride and anhydrous ethyl acetate).
  • the mixture is stirred at room temperature for 5 minutes and then cooled to -78 °C.
  • Trifluoromethanesulfonic anhydride (4 equivalents) is slowly added and the system is kept at -70 °C for 1 hour.
  • the reaction mixture is then kept at -45 °C to -40 °C for 3-16 hours, and quenched with a mixture of methanol and diisopropylethylamine.
  • the reaction mixture is allowed to warm to room temperature and the resin is washed with DMF (3x), tetrahydrofuran (2x), methanol (2x), and methylene chloride (2x).
  • a sample of the resin is then cleaved with a 30% cocktail of trifluoroacetic acid and methylene chloride for half an hour. The supernatant is evaporated to dryness and the residue is dissolved and analyzed by HPLC and LC-MS.
  • Glycosyl donor (6) is coupled with glycosyl acceptors (13), (14) and (15) to obtain the corresponding ⁇ -linked disaccharide (16), (17) and (18) respectively.
  • the trifluoroacetamido group brings extra flexibility to the sulfoxide glycosylation.
  • the sulfoxides are reactive and glycosylate unreactive nucleophiles such as glycosyl acceptor (13).
  • the high reactivity of glycosyl donors such as (6) and (12) can be appreciated by comparing them to other studied sulfoxides.
  • glycosyl acceptors (13) at >90% conversion can be achieved with 4 equivalents of glycosyl donors (6) or (12).
  • sulfoxides are used, up to 8 equivalents of sulfoxides have to be used to achieve the same conversion level.
  • Glycosyl donor (6) has been used to successfully glycosylate acceptors (13), (14) and (15), as shown in Figure 3.
  • the glycosylated products (16), (17) and (18) ( Figure 4) are obtained in >90% yield (as determined by cleaving the product from the resin with a TFA-CH Cl cocktail and analyzing it by HPLC).
  • combinatorial libraries are designed around 16 and 17.
  • the disaccharide core is derivatized with 8 different isocyanates and 12 different carboxylic acids (Figure 8).
  • the anomeric group of the acceptor sugar is either a ⁇ -thiophenyl group or a ⁇ , ⁇ -hydroxy group (lactols).
  • disaccharide core is derivatized with 6 different isocyanates and 8 different carboxylic acids, yielding a 48-member combinatorial library (Figure 9).
  • each disaccharide library the corresponding disaccharide immobilized on Rink Amide resin (16 or 17) is fully deprotected by treatment with LiOH in 1:1 THF-MeOH.
  • the deprotected resin is then suspended in a 4:1 mixture of methylene chloride and tetrahydrofuran, and aliquots of this suspension are dispensed into a Irori MicroKanTM containing an RF microtag.
  • the aliquots are calculated so that each MicroKanTM contains 15 mg resin (in the case of 16) or 20 mg resin (in the case of 17).
  • Each MicroKanTM and its RF tag are scanned into the Irori synthesis software and assigned an identification number.
  • the libraries are then synthesized according to the reaction schemes shown in Figures 10 and 11.
  • the results of LC-MS analyses are consistent with the production of the desired library compounds on the basis of their molecular weights. (See, Figures 14 and 15, below.)
  • the MicroKanTM containers containing the derivatized resins, are placed in separate test tubes and treated with a 30% TFA-CH Cl cocktail for 30 minutes. The supernatants are then transferred to a well of a microtiter plate and concentrated under vacuum using a Savant evaporator. The resulting residues are then reconstituted in 1 ml of DMSO and the solutions are aliquoted for control by LC-MS analysis, antibacterial screens and compound storage.
  • the LC-MS trace for a representative product is shown in Figure 12.
  • the analytical results obtained from the LC-MS analysis of this library are summarized in Figure 13.

Abstract

La présente invention concerne un procédé de synthèse de β-oligosaccharides, dans lequel on utilise des alkylsulfényl- ou arylsulfényl-2-déoxy-2-N-trifluoracétamidoglycopyranoses comme donneurs de glycosyle via la glycosylation du sulfoxyde, à la fois en solution et en phase solide. Une fois activés dans les conditions de glycosylation, ces donneurs produisent les β-glycosides de façon exclusive et avec un rendement élevé. Etant donné que le groupe trifluoroacétamido est facilement éliminé dans des conditions douces, le groupe amino correspondant peut être dérivatisé de manière adéquate, même en présence de groupes hydroxyle non protégés. L'invention concerne la conception, la construction et l'analyse de bibliothèques de disaccharides, ainsi qu'un procédé permettant de synthétiser le donneur de glycosyle.
PCT/US1999/002180 1998-02-03 1999-02-03 GLYCOSYLATION DE SULFOXYDE EN SOLUTION ET EN PHASE SOLIDE: SYNTHESE D'OLIGOSACCHARIDES A LIAISON β A L'AIDE DE DONNEURS 2-DEOXY-2-N-TRIFLUOROACETAMIDO-GLYCOPYRANOSYLE WO1999039201A1 (fr)

Priority Applications (4)

Application Number Priority Date Filing Date Title
EP99905611A EP1053471A1 (fr) 1998-02-03 1999-02-03 Glycosylation de sulfoxyde en solution et en phase solide: synthese d'oligosaccharides a liaison beta a l'aide de donneurs 2-deoxy-2-n-trifluoroacetamido-glycopyranosyle
CA002319339A CA2319339A1 (fr) 1998-02-03 1999-02-03 Glycosylation de sulfoxyde en solution et en phase solide: synthese d'oligosaccharides a liaison .beta. a l'aide de donneurs 2-deoxy-2-n-trifluoroacetamido-glycopyranosyle
AU25736/99A AU2573699A (en) 1998-02-03 1999-02-03 Solution and solid phase sulfoxide glycosylation: synthesis of beta-linked oligosaccharides using 2-deoxy-2-n-trifluoroacetamido-glycopyranosyl donors
JP2000529604A JP2002501932A (ja) 1998-02-03 1999-02-03 液相および固相スルホキシドグリコシル化：２−デオキシ−２−Ｎ−トリフルオロアセトアミド−グリコピラノシル供与体を使用したβ−連結オリゴ糖の合成

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US7357098P 1998-02-03 1998-02-03
US60/073,570 1998-02-03

Publications (1)

Publication Number Publication Date
WO1999039201A1 true WO1999039201A1 (fr) 1999-08-05

Family

ID=22114503

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1999/002180 WO1999039201A1 (fr) 1998-02-03 1999-02-03 GLYCOSYLATION DE SULFOXYDE EN SOLUTION ET EN PHASE SOLIDE: SYNTHESE D'OLIGOSACCHARIDES A LIAISON β A L'AIDE DE DONNEURS 2-DEOXY-2-N-TRIFLUOROACETAMIDO-GLYCOPYRANOSYLE

Country Status (5)

Country Link
EP (1) EP1053471A1 (fr)
JP (1) JP2002501932A (fr)
AU (1) AU2573699A (fr)
CA (1) CA2319339A1 (fr)
WO (1) WO1999039201A1 (fr)

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
COUTANT C., JACQUINET J.-C.: "2-DEOXY-2-TRICHLOROACETAMIDO-D-GLUCOPYRANOSE DERIVATIVES IN OLIGOSACCHARIDE SYNTHESIS: FROM HYALURONIC ACID TO CHONDROITIN 4-SULFATE TRISACCHARIDES.", JOURNAL OF THE CHEMICAL SOCIETY, PERKIN TRANSACTIONS 1, ROYAL SOCIETY OF CHEMISTRY, GB, 1 January 1995 (1995-01-01), GB, pages 1573 - 1581., XP002919524, ISSN: 0300-922X, DOI: 10.1039/p19950001573 *
LEZNOFF C. C.: "THE USE OF INSOLUBLE POLYMER SUPPORTS IN GENERAL ORGANIC SYNTHESIS.", JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, AMERICAN CHEMICAL SOCIETY, US, 1 January 1978 (1978-01-01), US, pages 327 - 333., XP002919525, ISSN: 0002-7863 *

Also Published As

Publication number Publication date
CA2319339A1 (fr) 1999-08-05
EP1053471A1 (fr) 2000-11-22
JP2002501932A (ja) 2002-01-22
AU2573699A (en) 1999-08-16

Similar Documents

Publication Publication Date Title
US5700916A (en) Solution and solid-phase formation of glycosidic linkages
Mathieux et al. Synthesis of glycopeptide sequences of repeating units of the mucins MUC2 and MUC 3 containing oligosaccharide side-chains with core 1, core 2, core 3, core 4 and core 6 structure
Merritt et al. n-Pentenyl mannoside precursors for synthesis of the nonamannan component of high mannose glycoproteins
De Meo et al. Chemical synthesis of glycosides of N-acetylneuraminic acid
US6462183B1 (en) Protected aminosugars
Lindberg et al. Efficient routes to glucosamine-myo-inositol derivatives, key building blocks in the synthesis of glycosylphosphatidylinositol anchor substances
Lu et al. 5-Azido neuraminic acid thioglycoside as sialylation donor
CA2757091A1 (fr) Synthese du sulfate d'heparane
US5635612A (en) Method of forming multiple glycosidic linkages in a single step
WO2000042057A1 (fr) Groupes protecteurs pour la synthese de glucides
Sawant et al. Formal synthesis of a disaccharide repeating unit (IdoA–GlcN) of heparin and heparan sulfate
US6040433A (en) Sulfinyl hexose derivatives useful for glycosylation
US20040019198A1 (en) Method of forming glycosidic bonds from thioglycosides using an N,N-dialkylsulfinamide
Alex et al. Synthesis of 2-azido-2-deoxy-and 2-acetamido-2-deoxy-D-manno derivatives as versatile building blocks
WO1999039201A1 (fr) GLYCOSYLATION DE SULFOXYDE EN SOLUTION ET EN PHASE SOLIDE: SYNTHESE D'OLIGOSACCHARIDES A LIAISON β A L'AIDE DE DONNEURS 2-DEOXY-2-N-TRIFLUOROACETAMIDO-GLYCOPYRANOSYLE
Jamoisa et al. How to improve chemical synthesis of laminaribiose on a large scale
US20050010044A1 (en) Polysaccharides and methods and intermediates useful for their preparation
Hinou et al. Synthesis of Amphiphilic Chitopentaose and Chitoheptaose Derivatives Using a Common Disaccharidic Synthon as the Chain Elongation Unit.
Traboni FEDERICO II
HUT67179A (en) Process for prepg. polymeric lewis-x saccharides
Nisic Stereoselective Synthesis of Glycosyl Amides by Traceless Staudinger Ligation of Glycosyl Azides
Bacherikov Strategy and preparation of some bulding blocks for synthesis of branched oligosaccarides
Pohl Synthesis of carbohydrate-based ligands for the selectins and galectins
Malik et al. Trends in Carbohydrate Research
Smoot Development of the arming participating group

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AL AM AT AU AZ BA BB BG BR BY CA CH CN CZ DE DK EE ES FI GB GE GH GM HR HU ID IL IN IS JP KE KG KR KZ LC LK LR LS LT LU LV MD MG MK MN MW MX NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT UA UG UZ VN YU ZW

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): GH GM KE LS MW SD SZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG

121 Ep: the epo has been informed by wipo that ep was designated in this application
DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
ENP Entry into the national phase

Ref document number: 2319339

Country of ref document: CA

Ref country code: CA

Ref document number: 2319339

Kind code of ref document: A

Format of ref document f/p: F

ENP Entry into the national phase

Ref country code: JP

Ref document number: 2000 529604

Kind code of ref document: A

Format of ref document f/p: F

NENP Non-entry into the national phase

Ref country code: KR

WWE Wipo information: entry into national phase

Ref document number: 1999905611

Country of ref document: EP

WWP Wipo information: published in national office

Ref document number: 1999905611

Country of ref document: EP

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

WWW Wipo information: withdrawn in national office

Ref document number: 1999905611

Country of ref document: EP