Inhibition of sialyltransferases of murine lymphocytes by disaccharide nucleosides.

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Abstract
Two disaccharide nucleosides, 5-fluoro-2', 3'-isopropylidene-5'-O-(4-N-acetamido-2, 4-dideoxy-3, 6, 7, 8-tetra-O-acetyl-1-methoxycarbonyl-D-glycero-α-D-galacto-octapyranosyl) uridine (KI-8110) and 2', 3'-di-O-acetyl-5'-O-(4-N-acetamido-2, 4-dideoxy-3, 6, 7, 8-tetra-O-acetyl-1-methoxycarbonyl-D-glycero-α-D-galacto-octapyranosyl) inosine (KI-8115) decreased in corporation of sialic acid into glycoconjugates on the murine lymphocyte surface. Since sialidase (EC 3.2.1.18) activity, CMP-NeuAc hydrolase (EC 3.1.4.40) activity and the incorporation of galactose or N-acetylgalactosamine into cell-surface glycoconjugates were all unaffected by these disaccharide nucleosides, the observed decrease in incorporation of sialic acid into glycoconjugates is considered to reflect the inhibition of sialyltransferase (EC 2.4.99.1) on the lymphocyte surface. KI-8110 and KI-8115 specifically inhibited the transfer of sialic acid to exogenous desialylated glycoproteins or exogenous desialylated glycolipids. Namely, KI-8100 and KI-8115 inhibited the transfer of sialic acid to desialylated fetuin, porcine submaxillary mucin (PSM) and bovine submaxillary mucin (BSM), all of which have Ser (Thr)-linked oligosaccharide chains, but did not inhibit transfer to desialylated α1-acid glycoprotein, which has only Asn-linked oligosaccharide chains. KI-8110 and KI-8115 inhibited the transfer of sialic acid to desialylated gangliosides GA1 and GA2, but not GA3. By using p-nitrophenyl-β-D-galactoside as an acceptor and cytidine 5'-monophosphate-[14C] N-acetylneuraminic acid (CMP-[14C] NeuAc) as a donor, two types of sialyl derivatives, α-sialosyl-(2→3)-p-nitrophenyl-β-D-galactoside and α-sialosyl-(2→6)-p-nitrophenyl-β-D-galactoside, were produced in murine lymphocytes. The production of α-sialosyl-(2→3)-p-nitrophenyl-β-D-galactoside was specifically inhibited by KI-8110 and KI-8115. KI-8110 and KI-8115 competitively inhibited sialic acid transfer by lymphocyte sialyltransferase to desialylated fetuin as an exogenous acceptor. The apparent K1 values were 2.3mM for KI-8110 and 2.5mM for KI-8115. When cytidine 5'-diphosphate (CDP), a known inhibitor of sialyltransferase, was incubated with lymphocyted for 24 h at 37°C, the inhibitory activity of CDP was dramatically decreased. However, such incubation did not affect the inhibitory activity of KI-8110 or KI-8115.