Materials and methods relating to glycosylation

摘要

Techniques to glycosylation are described, and more particularly to the production of glycosylation structures that are resistant to enzymatic degradation, thereby modulating one or more of their biological properties or those of therapeutic moieties incorporating them, and in particular to reacting activated carbohydrate substrates containing fluorine, such as 3-fluoro sialic acid compounds, with sugar acceptors to produce covalent conjugates of the sugar acceptor and one or more of the sialic acid compounds.

主权项

1. A method for glycosylating a therapeutic polypeptide, by forming a covalent conjugate between a 3-fluorosialic acid compound and a sugar acceptor covalently linked to the therapeutic polypeptide, said sugar acceptor including a terminal glycosyl residue, the method comprising contacting the sugar acceptor and 3-fluorosialic acid compound, the contacting step taking place under conditions suitable for reacting and covalently bonding the 3-fluorosialic acid compound to the sugar acceptor and yielding a glycosylation structure wherein said 3-fluorosialic compound is a terminal glycosyl group of said glycosylation structure, wherein the 3-fluorosialic acid compound does not comprise a cytosine monophosphate (CMP) group, and wherein the 3-fluorosialic acid compound is represented by general formula (I): wherein: Y1 is selected from —O—, —S—, or —NR—, wherein R is independently selected from H, C1-7 alkyl, C3-10 heterocyclyl, or C5-20 aryl; R1 is a leaving group effective to support and stabilize a negative charge, with the proviso that said leaving group is not a cytosine monophosphate (CMP) group; X1 is —CO2R, wherein R is as defined above; R2 is selected from H, halide or OH; R3 and R4 are each independently selected from H, —OR, —NR2 or —Z1(CH2)mZ2, where R is as defined above, Z1 is selected from —O—, —NR—, —CR2— and —S—, m is from 0 to 5 and Z2 is selected from —OR, —NR2 or —CN; with the proviso that R3 and R4 cannot both be H; R5 is H; R6 is selected from C1-7 alkyl; C1-7 hydroxyalkyl, C1-7 amino alkyl or C1-7 thioalkyl; R7 is represented by the formula: wherein Y2 is selected from N, O, S, and CH; Z3 is selected from H, hydroxyl, halide, C1-7 alkyl, C1-7 aminoalkyl, C1-7 hydroxyalkyl, or C1-7 thioalkyl; R9 and R10 are independently selected from H, hydroxyl, C1-7 hydroxyalkyl, C1-7 alkyl, C5-20 aryl, C(O)Z4, wherein Z4 is selected from C1-7 alkyl or C5-20 aryl, with the proviso that if Y2 is O or S, R10 is absent; or wherein R4 is other than hydroxyl, R7 may additionally be C1-7 hydroxyalkyl; R8 is hydrogen; or an oligomer of two or more molecules of formula (I); or stereoisomeric forms, tautomeric forms, salts, solvates, or chemically protected forms thereof.