Selective protection of mitogenically stimulated human lymphocytes but notleukemic cells from cytosine arabinoside-induced apoptosis by LY294002, a phosphoinositol-3 kinase inhibitor

Most malignant tumors have a defect in the Rb pathway that allows them to p roliferate autonomously, unrestricted by the availability of mitogens and g rowth factors. This defect preconditions tumor cells to be less sensitive t o inhibitors of growth factor receptors and/or signal transduction pathways that arrest normal cells, predominantly in G(0/1). Strategies were propose d, therefore, to combine cytotoxic agents with such inhibitors in order to selectively arrest normal host cells in a relatively resistant part of the cycle and thereby protect them during chemotherapy with agents targeting pr oliferating cells. The present study was designed to explore whether inhibi tion of phosphatidylinositol-3 kinase (P13K), one of the pivotal kinases in volved in signal transduction essential for cell proliferation can be consi dered in these strategies. Indeed, proliferation of phytohemagglutinin stim ulated human lymphocytes was prevented at 5 muM concentration of LY294002 ( LY), a P13K inhibitor, whereas several-times higher (greater than or equal to 20 muM) concentrations of LY were needed to affect proliferation of leuk emic HL-60, Molt-4, or Jurkat cells. LY prevented the exit of lymphocytes f rom G(0) concomitant with the suppression of induction of cyclins D2, D3 an d E, and phosphorylation of pRb. G(0/1) lymphocytes, that were initially st imulated in the absence of LY, were also inhibited from proliferating follo wing exposure to LY. The arrest of lymphocytes by LY was reversible and, af ter its removal, the cells asynchronously re-entered the cell cycle. LY, at a concentration of 5-20 muM, protected lymphocytes from apoptosis induced by ara-C but offered no protection at all to Jurkat cells treated under ide ntical conditions. The data suggest that inhibitors of P13K such as LY may be considered in strategies designed to shield normal cells from the cytoto xicity of chemotherapy by transiently arresting them in the cycle. An addit ional advantage of LY was the suppression of the protein level and activity of PKB/Akt, a kinase that, through the phosphorylation of the proapoptotic molecule BAD, protects cells from apoptosis. Because this LY-induced suppr ession was stronger in Jurkat cells (> 70%) than in lymphocytes (20%), the proapoptotic effects of PKB/Akt down-regulation appear to be selective towa rds tumor cells.