MODIFICATION AT C2 OF MYOINOSITOL 1,4,5-TRISPHOSPHATE PRODUCES INOSITOL TRISPHOSPHATES AND TETRAKISPHOSPHATES WITH POTENT BIOLOGICAL-ACTIVITIES

Novel 2-position-modified D-myo-inositol 1,4,5-trisphosphate [Ins(1,4, 5)P-3] analogues, DL-2-deoxy-2-fluoro-myo-inositol 1,4,5-trisphosphate [DL-2F-Ins(1,4,5)P-3], DL-myo-inositol 1,2,4,5-tetrakisphosphate [DL- Ins(1,2,4,5)P-4], DL-scyllo-inositol 1,2,4-trisphosphate [DL-sc-Ins(1, 2,4)P-3], scyllo-inositol 1,2,4,5-tetrakisphosphate [sc-Ins(1,2,4,5)P- 4] and scyllo-inositol 1,2,4,5-tetrakisphosphorothioate [sc-Ins(1,2,3, 5)PS4] were investigated for their ability to bind to the Ins(1,4,5)P- 3 receptor, mobilise intracellular Ca2+ stores and interact with metab olic enzymes. With the exception of sc-Ins(1,2,4,5)PS4, all the Ins(1, 4,5)P-3 analogues potently displaced [H-3]Ins(1,4,5)P-3 from its recep tor in bovine adrenal cortex and were apparently potent full agonists at the Ca2+ mobilising Ins(1,4,5)P-3 receptor of SH-SY5Y cells, giving respective IC50 and EC(50) values of: sc-Ins(1,2,4,5)P-4 (IC50 14 nM, EC(50) 77 nM), DL-2F-Ins(1,4,5)P-3 (IC50 25 nM, EC(50) 105 nM), DL-In s(1,2,4,5)P-4 (IC50 26 nM, EC(50) 163 nM), DL-sc-Ins(1,2,4)P-3 (IC50 5 2 nM, EC(50) 171 nM), compared to Ins(1,4,5)P-3 (IC50 4 nM, EC(50) 52 nM). sc-Ins(1,2,4,5)P-4 was equipotent to Ins(1,4,5)P-3 for Ca2+ relea se making it the most potent inositol tetrakisphosphate and indeed Ins (1,4,5)P-3 analogue yet characterised. In contrast, although sc-Ins(1, 2,4,5)PS4 (IC50 425 nM, EC(50) 1603 nM) was a significantly weaker lig and and agonist than Ins(1,4,5)P-3, it was a partial agonist of high i ntrinsic activity with maximally effective concentrations releasing on ly about 80% of Ins(1,4,5)P-3-sensitive Ca2+ stores of SH-SY5Y cells. Ins(1,4,5)P-3 and sc-Ins(1,2,4,5)P-4 were readily metabolised by Ins(1 ,4,5)P-3 3-kinase and 5-phosphatase activities, DL-2F-Ins(1,4,5)P-3 an d DL-sc-Ins(1,2,4)P-3 were resistant to 5-phosphatase, while sc-Ins(1, 2,4,5)PS4 and DL-Ins(1,2,4,5)P-4 were resistant to both 3-kinase and 5 -phosphatase activity and were potent inhibitors of the 5-phosphatase enzyme (K-i = 300 nM and 2.9 mu M, respectively). These results demons trate that modification of the 2-position of Ins(1,4,5)P-3, even with an anionic group, does not critically affect Ins(1,4,5)P-3 receptor bi nding interaction or Ca2+ release, suggesting that the 2-OH of Ins(1,4 ,5)P-3 fails to interact significantly with the binding site of its re ceptor. However, modification remote from the crucial vicinal 4,5-bisp hosphate can affect analogue efficacy in Ca2+ release.