2-DEOXY DERIVATIVE IS A PARTIAL AGONIST OF THE INTRACELLULAR MESSENGER INOSITOL 3,4,5,6-TETRAKISPHOSPHATE IN THE EPITHELIAL-CELL LINE T-84

We have synthesized the first deoxy analogues of myo-inositol 3,4,5,6- tetrakisphosphate (1) [Ins(3,4,5,6)P-4], rac-2-deoxy-myo-inositol 3,4, 5,6-tetrakisphosphate (rac-2), 2-deoxy-myo-inositol 1,4,5,6-tetrakisph osphate (ent-2), and rac-1-deoxy-myo-inositol 3,4,5,6-tetrakisphosphat e (rac-3). In order to evaluate the binding properties of the three de rivatives to the yet unidentified intracellular binding sites for Ins( 3,4,5,6)P4, the analogues were converted to membrane-permeant derivati ves. Starting with common inositol precursors, various forms of Barton -McCombie deoxygenation and classical protection/deprotection procedur es yielded the desired precursors rac-1-O-butyryl-2-deoxy-myo-inositol (rac-12), ent-3-O-butyryl-2-deoxy-myo-inositol (ent-12), and rac-2-O- butyryl-1-deoxy-myo-inositol (rac-19), respectively. Phosphorylation a nd subsequent deprotection yielded rac-2, ent-2, and rac-3. Alternativ ely, phosphorylation followed by alkylation with acetoxymethyl bromide gave the membrane-permeant derivatives 1-O-butyryl-2-deoxy-myo-inosit ol 3,4,5,6-tetrakisphosphate octakis(acetoxymethyl) ester (rac-5), 3-O -butyryl-2-deoxy-myo-inositol 1,4,5,6-tetrakisphosphate octakis(acetox ymethyl) eater (ent-5), and 2-O-butyryl-1-deoxy-myo-inositol 3,4,5,6-t etrakisphosphate octakis(acetoxymethyl) ester (rac-6), respectively. W e examined the potency of the membrane-permeant deoxy derivatives in i nhibition of calcium-mediated chloride secretion (CaMCS) in intact T-8 4 cells. Compared to the 1,2-di-O-butyryl-myo-inositol 3,4,5,6-tetraki sphosphate octakis(acetoxymethyl) ester (4), the membrane-permeant der ivative of Ins(3,4,5,6)P-4 (1), the 2-deoxy derivative (rac-5) exhibit ed a slightly weaker inhibitory effect, while the enantiomerically pur e 2-deoxy-Ins(1,4,5,6)P-4 (ent-5) and the l-deoxy derivative (rac-6) w ere inactive. As expected, the effect was stereoselective. Thus, the l -hydroxyl group is apparently essential for binding and the inhibitory effect of Ins(3,4,5,6)P4 on chloride secretion, whereas the 2-hydroxy l group plays a less important role.