Protected aminosugars

Details Protected aminosugars Protected aminosugars

1999-08-26

2002-10-08

Wilson, James O. (Department: 1623)

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

C536S018500, C536S124000

Reexamination Certificate

active

06462183

ABSTRACT:

This invention relates to methods for synthesis of oligosaccharides, especially those oligosaccharides which comprise amino sugar residues. In particular the invention relates to methods for solution phase, solid phase or combinatorial synthesis of oligosaccharides.
BACKGROUND OF THE INVENTION
Aminosugars are important constituents of various glycoconjugates (Schmidt and Kinzy, 1994). Examples include peptidoglycans, mucopolysaccharides, glycopeptides and proteins, oligosaccharides of human milk, and blood group determinants. They are often also encountered in bacterial and tumour-associated carbohydrate antigens, predominantly in the N-acetylated form or N-acylated with an aspartic acid residue (Toyokuni and Singhal, 1995). It is therefore evident that these biological glycoconjugates are of immense interest to the medicinal chemist, and therefore that there is a great need in the art to be able to synthesise these compounds in a facile and cost-effective manner.
Oligosaccharide synthesis using aminosugars requires the presence of a suitable amino protecting group. A number of protecting groups have been proposed, but so far all of the agents which are available suffer from serious disadvantages. For example, glycosylation with donors derived from 2-N-acetyl protected aminosugars proceeds via neighbouring group participation; however, formation of the relatively stable oxazoline intermediate dramatically reduces the overall speed and yield of the reaction (Zurabyan et al, 1994). Therefore, various 2-deoxy-2-aminosugar donors, displaying the neighbouring group activity described, but lacking the ability to form stable oxazolines, have been developed; the most widely used of these are the phthalimido protected monomers (Sasaki et al, 1978). The phthalimide group participates strongly during glycoside formation and gives excellent stereocontrol of the 1,2-trans-glycoside product (Lemieux et al, 1982), furthermore the aminosugar donors do not form stable orthoamides (Lemieux et al, 1982) and cannot form oxazolines. The major disadvantage of using the phthalimide group lies in the vigorous conditions required for its removal, namely heating with methanolic hydrazine, which often results in partial product decomposition. Strongly basic conditions are also required for the removal of the N-sulfonyl (Griffith and Danishefsky, 1990) and N-haloacetyl protecting groups (Shapiro et al, 1967), resulting in similar problems.
The allyloxycarbonyl (Alloc) protected amino sugar donors display a similar activity to their phthalimide counterparts when employed under Lewis acid-catalysed conditions. However, the Alloc group has the advantage that it can be removed under extremely mild conditions, using tetrakis(triphenylphosphine)palladium in the presence of a mild base (Hayakawa et al, 1986). The major disadvantage associated with the Alloc group lies in its ability to form a stable oxazolidinone intermediate, which in the presence of unreactive acceptors tends to remain as the major product, and reduces the speed and yield of the reaction (Boullanger et al, 1987). 2,2,2-Trichloroethyl-protected aminosugars contain a strongly participating group that, unlike phthalimide, does not deactivate adjacent hydroxyl groups which may subsequently be required as glycosyl acceptors. They can be removed under relatively mild and selective conditions, using zinc and acetic acid, and do not form oxazoline intermediates during glycosylation. However, this protecting group has the disadvantage that benzyl groups cannot be introduced without premature loss of the protecting group as well (Imoto et al, 1987).
Tetrachlorophthaloyl-protected aminosugar donors have been demonstrated to afford high yields of 1,2-trans-glycosides (Castro-Palomino and Schmidt, 1995), even in the presence of poorly reactive acceptors. Once more, however, the NaBH
4
-mediated deprotection is the limiting factor for this particular protecting group.
The azide group has received much attention in aminosugar chemistry, since it serves as a masked, non-participating amino functionality, thereby allowing the synthesis of 1,2-cis-linked 2-amino-2-deoxy glycosides (Palsen, 1982). However the preparation of 2-azido-2-deoxy sugars is protracted, costly, and often dangerous, using either azidonitration (Lemieux and Ratcliffe, 1979), diazo-transfer reactions (Buskas et al, 1994), azidochlorination (Bovin et al, 1986), nitrosation of N-benzyl derivatives (Dasgupta and Garegg, 1989) or reactions of 1,6-anhydrosugars (Tailler et al, 1991 and Paulsen and Stenzel, 1978).
Other non-participating protecting groups that have been reported are 2,4-dinitrophenyl (Kaifu and Osawa, 1977) and p-methoxybenzylimino (Mootoo and Fraser-Reid, 1989), both of which are complicated to introduce and require harsh deprotection conditions which result in loss of product.
A hydrazine-labile primary amino-protecting group, N-1-(4,4-dimethyl-2,6-dioxocyclohexylidene)ethyl (Dde), has been reported for protection of lysine side chains during SPPS (Bycroft et al, 1993). This group was modified for use as a carboxy-protecting group in SPPS when the 2-(3-methylbutyryl)dimedone analogue of 2-acetyldimedone was condensed with 4-aminobenzylalcohol to afford 4-{N-[1-(4,4-dimethyl-2,6-dioxocyclo-hexylidene)-3-methylbutyl]-amino}benzyl ester (ODmab) (Chan et al, 1995). These two protecting groups were reported to be stable to the Fmoc deprotecting conditions widely used in solid phase peptide synthesis (SPPS), ie 20% piperidine in dimethylformamide (DMF).
Dde has been widely used in the field of SPPS as an orthogonal amino protecting group to the well established Fmoc/t-Boc methodology (Fields and Noble, 1990). Until now its use has remained within this area, and therefore its use as a protecting group in the field of carbohydrate chemistry is novel. In particular, the use of Dde or ODMab in oligosaccharide synthesis has not been suggested.
We have now surprisingly found that Dde can be used as a non-participating amino sugar protecting group, which can be introduced and removed in a facile and cost-effective manner. We have shown that the vinylogous amide protection afforded by the Dde type group is achieved by simply refluxing the unprotected amino sugar with the precursor, eg. 2-acetyldimedone in the case of Dde, in anhydrous ethanol. Using a Dde-protected aminosugar, we have performed a variety of chemical modifications upon the protected molecule in order to demonstrate the stability of this vinylogous amide type protection towards commonly encountered reactions involved in carbohydrate modification.
SUMMARY OF THE INVENTION
In one aspect, the invention provides a compound useful as a reagent for solution and/or solid phase synthesis of sugar-containing compounds, comprising a sugar carrying one or more primary amine groups protected with a 2-substituted-1,3-dioxo compound of General Formula I or General Formula II:
in which
R
1
and R
2
may be the same or different, and is each hydrogen or C
1-4
alkyl,
R′ is an amino sugar, a glycosylamine, or an oligosaccharide comprising at least one aminosugar or one glycosylamine unit, in which the sugar is coupled via an amino group,
and R″ is alkyl, substituted alkyl, aryl, substituted aryl, cycloalkyl or substituted cycloalkyl.
Any sugar or oligosaccharide bearing an amino group may be used.
In a preferred embodiment, the invention provides a reagent for solution phase synthesis of sugar-containing compounds, comprising a cyclic 2-substituted-1,3-dioxo compound of General Formula I or II as defined above, in which R′ is as defined above.
The compounds of the invention are suitable for use in methods of solid-phase oligosaccharide synthesis, in which sugar units are covalently linked to a resin. Any suitable linker compound may be used. For example, the covalent linkage to the resin may suitably be provided by a —CONH—, —O—, —S—, —COO—, —CH═N—, —NHCONH—, —NHCSNH, or —NHNH— grouping, eg. Spacer— CONH-resin, Spacer-O-resin, Spacer-S-resin, Spacer-CO
2
-resin, Spacer-CH═N-resin, Spacer-NHCON

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