DISACCHARIDE POLYPHOSPHATES BASED UPON ADENOPHOSTIN-A ACTIVATE HEPATIC D-MYO-INOSITOL 1,4,5-TRISPHOSPHATE RECEPTORS

The glyconucleotides adenophostin A and B are the most potent known ag onists at type 1 inositol trisphosphate [Ins(1,4,5)P-3] receptors, alt hough their stuctures differ markedly from that of Ins(1,4,5)P-3. Equi librium competition binding with [H-3]Ins(1,4,5)P-3 and unidirectional Ca-45(2+) flux measurements were used to examine the effects of adeno phostin A in hepatocytes, which express predominantly type 2 Ins(1,4,5 )P-3 receptors. Both Ins(1,4,5)P-3 (K-d = 8.65 +/- 0.98 nM) and adenop hostin A (K-d = 0.87 +/- 0.20 nM) bound to a single class of [H-3]Ins( 1,4,5)P-3-binding site and each fully mobilized the same intracellular Ca2+ pool; although, adenophostin A (EC50 = 10.9 +/- 0.7 nM) was more potent than Ins(1,4,5)P-3 (EC50 = 153 +/- 11 nM). Working on the assu mption that it is the phosphorylated glucose component of the adenopho stins that mimics the critical features of Ins(1,4,5)P-3, we synthesiz ed various phosphorylated disaccharide analogs containing this structu re. The novel disaccharide-based analogs, sucrose 3,4,3'-trisphosphate [Sucr(3,4,3')P-3], alpha,alpha'-trehalose 3,4,3',4'-tetrakisphosphate [Trehal(3,4,3',4')P-4], alpha,alpha'-trehalose 2,4,3',4'-tetrakisphos phate [Trehal(2,4,3',4')P-4], and methyl -O-((alpha-D-glucopyranosyl)- beta-D-ribofuranoside 2,3',4'-trisphosphate [Rib(2,3',4')P-3], were al l able to mobilize the same intracellular Ca2+ pool as Ins(1,4,5)P-3 a nd adenophostin A; although, none was as potent as adenophostin A. The rank order of potency of the analogs, adenophostin A > Ins(1,4,5)P-3 approximate to Rib(2,3',4')P-3 > Trehal(2,4,3',4')P-4 > Glc(2',3,4)P-3 approximate to Trehal(3,4,3',4')P-4 > Sucr(3,4,3')P-3, was the same i n radioligand binding and functional assays of hepatic Ins(1,4,5)P-3 r eceptors. Both Rib(2,3',4')P-3, which was as potent as Ins(1,4,5)P-3, and Trehal(2,4,3',4')P-4 bound with significantly higher affinity (sim ilar to 27 and similar to 3-fold, respectively) than the only active c arbohydrate agonist of Ins(1,4,5)P-3 receptors previously examined [Gl c(2',3,4)P-3]. We conclude that phosphorylated disaccharides provide n ovel means of developing high-affinity ligands of Ins(1,4,5)P-3 recept ors.