Inhibition of protein kinase C by ether-linked lipids is not correlated with their antineoplastic activity on WEHI-3B and R6X-B15 cells

Abstract

To test the hypothesis that the action of antineoplastic ether-linked lipids in leukemic cells is associated with their ability to inhibit protein kinase C (PKC), we have compared the effects of two ether-linked lipids. 1- O-hexadecyl-2- O-methyl- sn-glycero-3-phosphocholine (ET 16-OCH 3-GPC) and 1-O- hexadecyl-2-O- methyl-sn- glycero-3-(S-β- d -1′- thioglucopyranosyl)-sn- glycerol (ET 16-OCH 3-β-thio-Glc), on two different leukemic cell lines (WEHI-3B and R6X-B15). ET 16-OCH 3-GPC killed WEHI-3B cells with an EC 50 value of 2.5 μM, whereas it was far less effective against R6X-B15 cells (EC 50 = 40 μM). In contrast, the β anomer of ET 16-OCH 3-ß-thio-Glc did not kill either cell line at concentrations up to 40 μM. Both ET 16-OCH 3-GPC and ET 16-OCH 3-thio-Glc inhibited 12- O-tetradecanoylphorbol 12,13-dibutyrate (TPA)-induced PKC translocation in both WEHI-3B and R6X-B15 cells. When WEHI-3B cells were first exposed to TPA, and then to ET 16-OCH 3-GPC, no significant decrease in PKC activity in the particulate fraction was noticed. When, however, the cells were first exposed to ET 16-OCH 3-GPC and then to TPA, the enzyme activity in the particulate fraction was decreased by 20–30%. A phorbol dibutyrate binding assay showed that the decrease in membrane-associated PKC activity and the increase in cytosolic PKC activity did not result from impeded enzyme translocation. These results suggest that the similar PKC inhibitory potency of ET 16-OCH 3-GPC and ET 16-OCH 3ß-thio-Glc: (a) is not correlated with the widely different cytotoxicities of these agents and (b) is probably due to interference with the binding of diacylglycerol/phosphatidylserine or TPA to PKC. Taken together, these results suggest that the ether-linked lipids compete with diacylglycerol/phosphatidylserine or TPA for binding sites on PKC required for enzyme activation.