Cytotoxic Ether Lipid Analogues Selective Inhibition of Phosphatidylinositol Phospholipase C by Updated Version

The ether lipid analogue l-octadecyl-2-methyl-rac-glycero-3-phosphocholine (ET-18-OCH.,) has been shown to be a direct inhibitor of Swiss 3T3 fibroblast and BG1 ovarian adenocarcinoma cell cytosolic phosphoinositide selective phospholipase C (PIPLC) using |3H|-phosphatidylinositol-(4, 5)-bisphosphate (|'II|I"II'<) as the substrate. The inhibition occurred when ET-18-OCH., was incorporated into the |3H|PIP2 substrate micelles, with 50% inhibition «!(.,„) occurring at a I I 1K-(K II,: |3H|PIP2 ratio of 0.04, or an assay concentration of 0.4 MM,and when ET-18-OCH., was added directly to the incubation, with an ICs>of 9.6 MM.Lipid prepared from cells exposed to cytotoxic concentrations of ET18-OCH, for 18 h also inhibited PIPLC with an 1C»< 1 MM.The noncytotoxic analogue 1-O-alkyl-l-hydroxy-sn-glycero^-phosphocholine inhibited PIPLC when incorporated into the |3H|PIP2 substrate micelles, but lipid from cells grown with 5 MMl-O-alkyl-2-hydroxy-Ã-/iglycero-3-phosphocholine did not inhibit PIPLC. BG1 cells, which were more sensitive than Swiss 3T3 fibroblasts to growth inhibition by ET18-OCH.,, had a cytosolic PIPLC activity one-third that of Swiss 3T3 cells. NIH 3T3 cells exhibited the same sensitivity to growth inhibition by ET-18-OCHj as Swiss 3T3 cells and had a similar level of PIPLC. v.v/.vNIH 3T3 cells were relatively resistant (>3-fold) to growth inhibition by ET-18-OCHj and had a cytosolic PIPLC activity more than twice that of the wild type cells. ET-18-OCH., was a weak inhibitor, K .„ >100 MM,of phospholipase D activity in NIH 3T3 cell membranes. In intact NIH 3T3 cells ET-18-OCH., at cytotoxic concentrations did not inhibit phospholipase D or phosphatidylcholine-selective phospholipase C activ ity. The results show that the ether lipid analogues at cytotoxic concen trations are selective inhibitors of PIPLC and that the inhibition of PIPLC may be related to the growth inhibitory activity of the ether lipid analogues.