Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1994-10-13
1997-05-06
Rollins, John W.
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
424 945, 435193, C07G 300, A61K 3851, C12N 910
Patent
active
056272715
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
This invention relates to glycolipids and to their preparation and use.
BACKGROUND OF THE INVENTION
Glycolipids are compounds of importance in therapy. In particular, glycosphingolipids are naturally-occurring compounds found in the membranes of all animal cells and in some plant cells. One such compound is cytolipin H, a pure lipid hapten, which has been isolated from human epidermoid carcinoma extracts.
Cytolipin H is N-lignoceroyl-1-sphingosyl lactoside. This is a compound of formula I (see formulae below) when Q is H, X is --NH--CO--C.sub.23 H.sub.47, Y is --CH.dbd.CH--C.sub.13 H.sub.27 and (sac).sub.m+n, is a lactose residue. A synthesis of such compounds is reported by Nicolaou et al, JACS (1988) 110: 7910-7912, and comprises coupling a protected di- or trisaccharide with a protected sphingosine azide which is then elaborated.
Yu et al, J. Biol. Chem. (1976) 251(1): 198-203, discloses that a sialyltransferase in mouse brain microsomes catalyses the synthesis of sialosylgalactosylceramide from added galactocerebroside and CMP-NANA. The product, i.e. NeuNAc(.alpha.,2-3)gal(.beta.,1-1) cer, and the substrate, are compounds of formulae I and II in which X is --NH--CO--C.sub.12 H.sub.25 and Y is --CH.dbd.CH--C.sub.13 H.sub.27. The enzymatic activity is apparently dependent on the ceramide structure of the glycolipids.
Products of the ceramide type, i.e. containing two hydrophobic residues, have serological activity that depends on the nature of Y; see Graf et al, Chemistry and Physics of Lipids (1974) 13: 367-371.
EP-A-0212400 discloses a chemical synthesis of compounds of formula II wherein n=1, sac is glucopyranose, X is --NH--CO--(C.sub.14-24 alkyl) and Y is --CH.dbd.CH--C.sub.13 H.sub.27 or an analogous group. Intermediates in this synthesis are of the same structure except that X is N.sub.3 or NH.sub.2. The latter is suitable for elaboration to a variety of groups X.
Zimmermann et al, J. Carbohydrate Chem. 7(2): 435-452 (1988), disclose similar compounds, and also analogous compounds wherein n=2. For example, in order to prepare lactosylceramide, the per-O-acetylated trichloroacetimidate of lactose is first coupled with a hydroxysphingosineazide;the product is reduced and elaborated.
SUMMARY OF THE INVENTION
The present invention is based on the discovery that a glycolipid of the type obtained by coupling a saccharide and a sphingosine, but not a ceramide, i.e. X is N.sub.3 or NH.sub.2, is a satisfactory substrate for an enzymatic saccharide coupling reaction.
The substrate has formula II (preferably the natural configuration shown below), i.e.
The product has formula I (preferably the natural configuration shown below), i.e.
Many of the products and substrates are new. The advantage of using the new procedure to make glycolipids having at least two sugar moieties is that, except for elaboration to give any desired group X, the more expensive enzymatic reactions are conducted after the chemical modifications, giving greater efficiency and higher yields, as well as preserving the desired stereochemistry. The elaborated products are known compounds, or may be new compounds having properties analogous or perhaps superior to the related known compounds.
DESCRIPTION OF THE INVENTION
In the starting glycolipid (II), n is preferably 1 or 2. m is usually one, so that the novel process is used to add one sugar moiety at a time. If a glycolipid (II) in which n=1 is used as starting material, and many such compounds can be prepared by generally known techniques, the process of the invention can be repeated as many times as necessary to give a glycolipid having the desired number of sugar moieties.
The enzyme that is used in the invention is typically of the type that is known for the biosynthesis of N-linked sugars in glycoproteins. It is surprising that such known enzymes will also utilise glycolipids (II) as a substrate but, as is evident from the data herein, that is the case. Suitable sialyltransferases (.alpha.2-3 and .alpha.2-6) may be obtained from porcine or rat liver, a
REFERENCES:
Zimmerman, P. et al. (1988) "Azidosphingosine Glycosylation in Glycosphingolipid Synthesis" J. Carbohydrate Chemistry, & (2):435-452.
Nicolaou, K. et al. (1988) "A Practical and Enantioselective Synthesis of Glycosphingolipids and Related Compounds. Total Synthesis of Globotriaosylceramide (GB3)" J. Am. Chem. Soc., 110:7910-7912.
Yu, R. et al. (1976) "In Vitro Biosynthesis of Sialosylgalacto-sylceramide (G7) by Mouse Brain Microsomes" The Journal of Biological Chemistry, 251(1):196-203.
Graf, L. et al. (1974) "Serological Activity of Glycosphingolipids: Effects of Chain Length of the Fatty acid Residue in Cytolipin H" Chemistry and Physics of Lipids, 13:367-371.
Flitsch Sabine L.
Guilbert Benedicte
Genzyme Limited
Gosz William G.
Rollins John W.
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