Suppression of esophageal tumor growth and chemoresistance by directly targeting the PI3K/AKT pathway

Esophageal cancer is the sixth most common cause of cancer-related deaths worldwide. Novel therapeutic intervention is urgently needed for this deadly disease. The functional role of PI3K/AKT pathway in esophageal cancer is little known. In this study, our results from 49 pairs of human esophageal tumor and normal specimens demonstrated that AKT was constitutively active in the majority (75.5%) of esophageal tumors compared with corresponding normal tissues. Inhibition of the PI3K/AKT pathway with specific inhibitors, wortmannin and {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002, significantly reduced Bcl-xL expression, induced caspase-3-dependent apoptosis, and repressed cell proliferation and tumor growth in vitro and in vivo without obvious toxic effects. Moreover, significantly higher expression level of p-AKT was observed in fluorouracil (5-FU)-resistant esophageal cancer cells. Inactivation of PI3K/AKT pathway markedly increased the sensitivity and even reversed acquired resistance of esophageal cancer cells to chemotherapeutic drugs in vitro. More importantly, the resistance of tumor xenografts derived from esophageal cancer cells with acquired 5-FU resistance to chemotherapeutic drugs was significantly abrogated by wortmannin treatment in animals. In summary, our data support PI3K/AKT as a valid therapeutic target and strongly suggest that PI3K/AKT inhibitors used in conjunction with conventional chemotherapy may be a potentially useful therapeutic strategy in treating esophageal cancer patients.     

Id-1 activation of PI3K/Akt/NFkappaB signaling pathway and its significance in promoting survival of esophageal cancer cells

Inhibitor of differentiation or DNA binding (Id-1) is a helix-loop-helix protein that is over-expressed in many types of cancer including esophageal cancer. This study aims to investigate its effects on the phosphatidylinositol-3-kinase (PI3K)/Akt/ nuclear factor kappa B (NFkappaB) signaling pathway and the significance in protecting esophageal cancer cells against apoptosis. We found elevated expression of phosphorylated forms of Akt, glycogen synthase kinase 3beta and inhibitor of kappa B, as well as increased nuclear translocation of NFkappaB subunit p65 and NFkappaB DNA-binding activity, in esophageal cancer cells with stable ectopic Id-1 expression. Transient transfection of Id-1 into HEK293 cells confirmed activation of PI3K/Akt/NFkappaB signaling and the effects were counteracted by the PI3K inhibitor LY294002. Treatment with tumor necrosis factor-alpha (TNF-alpha) elicited a significantly weaker apoptotic response, following a marked and sustained activation of Akt and NFkappaB in the Id-1-over-expressing cells, compared with the vector control. The effects of Id-1 on the PI3K/Akt/NFkappaB signaling pathway and apoptosis were reversed in esophageal cancer cells transfected with siRNA against Id-1. In addition, inhibition of PI3K or NFkappaB signaling using the PI3K inhibitor LY294002 or the NFkappaB inhibitor Bay11-7082 increased the sensitivity of Id-1-over-expressing esophageal cancer cells to TNF-alpha-induced apoptosis. Our results provide the first evidence that Id-1 induces the activation of PI3K/Akt/NFkappaB signaling pathway, and protects esophageal cancer cells from TNF-alpha-induced apoptosis in vitro. Inactivation of Id-1 may provide us with a novel strategy to improve the treatment and survival of patients with esophageal cancer.     

PTEN and phosphatidylinositol 3'-kinase inhibitors up-regulate p53 and block tumor-induced angiogenesis: evidence for an effect on the tumor and endothelial compartment

Previous work from our laboratory demonstrated that PTEN regulates tumor-induced angiogenesis and thrombospondin 1 expression in malignant glioma. Herein, we demonstrated the first evidence that the systemic administration of a phosphatidylinositol 3'-kinase (PI3K) inhibitor (LY294002) has antitumor and antiangiogenic activity in vivo. We show that PTEN reconstitution diminished phosphorylation of AKT, induced the transactivation of p53 (7.5-fold induction) and increased the expression of p53 target genes, p21(waf-1) and insulin-like growth factor binding protein 3 in glioma cells. PTEN and LY294002 induced p53 activity in human brain endothelial cells, suggesting that PTEN and PI3K pathways can suppress the progression of cancer through direct actions on tumor and endothelial cells. The capacity of PTEN and LY294002 to inhibit U87MG or U373MG glioma growth was tested in an ectopic skin and orthotopic brain tumor model. LY294002 inhibited glioma tumor growth in vivo, induced tumor regression, decreased the incidence of brain tumors, and blocked the tumor-induced angiogenic response of U87MG cells in vivo. These data provide evidence that both PTEN and PI3K inhibitors regulate p53 function and display in vivo antiangiogenic and antitumor activity. These results provide evidence that the two tumor suppressor genes, PTEN and p53, act together to block tumor progression in vivo. Our data provide the first preclinical evidence for the in vivo efficacy for LY294002 in the treatment of malignant gliomas.     

Inhibition of EGFR/PI3K/AKT cell survival pathway promotes TSA's effect on cell death and migration in human ovarian cancer cells

Trichostatin A (TSA), a hydroxamate-type inhibitor of mammalian histone deacetylases, is emerging as one of a potentially new class of anticancer agents. TSA is known to act by promoting the acetylation of histones, leading to uncoiling of chromatin and activation of a variety of genes implicated in the regulation of cell survival, proliferation, differentiation, and apoptosis. In addition, there is an increasing appreciation of the fact that TSA may act through mechanisms other than induction of histone acetylation. Accumulated experimental data indicate that TSA activates phosphatidyl inositol-3-kinase (PI3K)/AKT signaling. Using human ovarian cancer cell line Caov3 cells, we observed that TSA induced cell death in a time- and dose-dependent manner and also inhibited cell migration. TSA transiently activated EGFR tyrosine phosphorylation and AKT activation in a time- and dose-dependent manner, which had been inhibited by EGFR inhibitor PD153035 and PI3 kinase inhibitor LY294002. We also observed that TSA transiently induced survivin expression that had been inhibited by PD153035 and LY294002, suggesting that TSA-induced survivin expression is mediated by EGFR/PI3 kinase pathway. Combination of EGFR inhibitor 153035 or PI3 kinase inhibitor LY294002 with TSA enhanced TSA-induced cell death and TSA reduction of cell migration. Collectively, our data demonstrate that TSA transiently activated EGFR/PI3K/AKT cell survival pathway, leading to expression of survivin. Inhibition of this pathway enhanced TSA-induced cell death and inhibited cell migration. Our data suggest that combination of EGFR/PI3K/AKT cell survival pathway inhibitors with TSA be a better approach to ovarian cancer treatment.     

Homotypic gap junctional communication associated with metastasis suppression increases with PKA activity and is unaffected by PI3K inhibition

Loss of gap junctional intercellular communication (GJIC) between cancer cells is a common characteristic of malignant transformation. This communication is mediated by connexin proteins that make up the functional units of gap junctions. Connexins are highly regulated at the protein level and phosphorylation events play a key role in their trafficking and degradation. The metastasis suppressor breast cancer metastasis suppressor 1 (BRMS1) upregulates GJIC and decreases phosphoinositide-3-kinase (PI3K) signaling. On the basis of these observations, we set out to determine whether there was a link between PI3K and GJIC in tumorigenic and metastatic cell lines. Treatment of cells with the well-known PI3K inhibitor LY294002, and its structural analogue LY303511, which does not inhibit PI3K, increased homotypic GJIC; however, we found the effect to be independent of PI3K/AKT inhibition. We show in multiple cancer cell lines of varying metastatic capability that GJIC can be restored without enforced expression of a connexin gene. In addition, while levels of connexin 43 remained unchanged, its relocalization from the cytosol to the plasma membrane was observed. Both LY294002 and LY303511 increased the activity of protein kinase A (PKA). Moreover, PKA blockade by the small molecule inhibitor H89 decreased the LY294002/LY303511-mediated increase in GJIC. Collectively, our findings show a connection between PKA activity and GJIC mediated by PI3K-independent mechanisms of LY294002 and LY303511. Manipulation of these signaling pathways could prove useful for antimetastatic therapy.     

Epithelial V‐like antigen 1 promotes hepatocellular carcinoma growth and metastasis via the ERBB‐PI3K‐AKT pathway

The role of epithelial V‐like antigen 1 (EVA1) has been well studied in thymic development and homostasis; however, its putative relationship with cancer remains largely unknown. Therefore, here we investigated the role of EVA1 in hepatocellular carcinoma. Interestingly, EVA1 expression was significantly increased in hepatocellular carcinoma (HCC) and was also associated with a poor prognosis and recurrence in HCC patients. Overexpression of EVA1 promoted cell growth, invasion and migration in vitro. Consistently, knockdown of EVA1 expression inhibited proliferation and migration in vitro, while repressing metastasis of HCC cells in vivo. RNA‐seq analysis indicated that EVA1 is able to upregulate the expression of genes in the ERBB3‐PI3K pathway. Accordingly, an increased level of AKT phosphorylation was detected in HCC cells after EVA1 overexpression. LY294002, a PI3K inhibitor, inhibited AKT phosphorylation and rescued the tumor‐promoting effect of EVA1 overexpression. Altogether, the present study has revealed the oncogenic role of EVA1 during HCC progression and metastasis through the ERBB‐PI3K‐AKT signaling pathway, reiterating the potential use of EVA1 as a therapeutic target and/or prognostic marker for HCC.     

PI3K/AKT信号通路阻断药联合小檗碱对前列腺癌DU145细胞生长的抑制作用

目的 探讨采用磷脂酰肌醇3-激酶(PI3K)抑制药——LY294002抑制磷脂酰肌醇-3-激酶/丝苏氨酸蛋白激酶(PI3K/AKT)信号通路联合小檗碱对前列腺癌DU145细胞生长的抑制作用.方法 采用MTT法检测细胞增殖情况;采用流式细胞仪检测细胞凋亡率和细胞周期分布情况;采用Western blot检测细胞中PI3K、p-AKT、cyclin D1、bcl-2、BAX蛋白表达情况.结果 随着小檗碱作用浓度的增加,DU145细胞的增殖率逐渐降低;不同浓度小檗碱与LY294002联合处理对DU145细胞增殖的抑制作用均较相同浓度小檗碱单独处理增强(P﹤0.05).与空白对照组相比,小檗碱组DU145细胞的凋亡率增高(P﹤0.05),G0/G1期细胞所占比例增高(P﹤0.05),PI3K、p-AKT、cyclin D1、bcl-2蛋白表达水平均下调(P﹤0.05),BAX蛋白表达水平上调(P﹤0.05).与小檗碱组和空白对照组相比,小檗碱+LY294002组DU145细胞的凋亡率增高(P﹤0.05),G0/G1期细胞所占比例增高(P﹤0.05),PI3K、p-AKT、cyclin D1、bcl-2蛋白表达水平均下调(P﹤0.05),BAX蛋白表达水平上调(P﹤0.05).结论 小檗碱能够抑制DU145细胞增殖,并促进细胞凋亡和细胞周期阻滞,采用LY294002阻断PI3K/AKT信号通路能够明显增强小檗碱对前列腺癌DU145细胞生长的抑制作用.     

Ad-PTEN增强LY294002对人胶质瘤细胞系U251的生长抑制作用

目的在体外实验中,利用携带野生型PTEN基因的重组腺病毒（Ad—PTEN）和P13K的小分子抑制剂LY294002联合抑制P13K／AKT信号通路,观察两者联合对人胶质瘤细胞系U251生长有无正向协同作用及探讨产生这种作用的机制。方法15251细胞按处理方式不同分为4组：DMSO对照组、空载病毒对照组、LY294002组和联合组（LY294002＋Ad—PTEN组）。在U251细胞系中导入LY294002并转染Ad—PTEN病毒后,提取总蛋白,用Westernblotting检测PTEN表达状态及P13K、AKT表达情况;用MTT法检测Ad—PTEN和LY294002对U251生长抑制率、流式细胞仪检测细胞周期、AnnexinV法检测细胞凋亡,观察导人LY294002和转染Ad—PTEN后U251增殖能力的改变;用划痕实验、Transwell实验观察LY294002及Ad—PTEN对U251迁移和侵袭能力的影响;Westernblotting检测P13K／AKT信号通路下游相关蛋白表达水平变化。结果Westernblotting显示：与DMSO组和空载病毒组相比,LY294002组P13K、AKT蛋白表达水平明显降低,联合组（LY294002＋Ad—PTEN）的WEN蛋白明显上调,而P13K、AKT蛋白下降水平比LY294002组更为显著。联合组与LY294002组、空载病毒组、DMSO组相比细胞周期阻滞在G0／G1期;联合组凋亡率比其他三组明显增加;自培养第2天起,联合组细胞增殖速率呈下降趋势。联合组侵袭和迁移细胞的相对数少于LY294002组、空载病毒组和DMSO组。Westernblotting结果证明位于P13K／AKT下游的MMP-2、MMP-9、PCNA、Bcl-2、CyclinD1、NF—KB、FAK蛋白表达水平显著下调。结论小分子抑制剂LY294002能够有效抑制U251中P13K、AKT蛋白的表达,联合转染Ad—PTEN后不仅能提高PTEN蛋白的表达水平,对P13K、AKT的抑制更为显著。在体外实验中,联合Ad—PTEN和LY294002能够有效地通过阻滞细胞周期、减少细胞凋亡来抑制U251的增殖能力,并能够抑制细胞的侵袭和迁移能力,两者联合使用具有正向协同作用。联合Ad—PTEN和LY294002对U251的生长与侵袭的抑制作用与P13K／AKT下游的MMP-2、MMP-9、PCNA、Bcl-2、CyclinD1、NF—KB、FAK蛋白表达水平下调有关。联合腺病毒转染技术和小分子抑制剂调控P13K／AKT信号通路是治疗胶质瘤的新途径。     

HER2/neu过表达通过PI3K/Akt通路对乳腺癌细胞MCF7中p53基因表达及细胞生长和放疗敏感性的影响

目的 研究HER2／neu基因过表达通过磷脂酰肌醇-3-激酶(PI3K)通路对乳腺癌细胞MCF7野生型p53基因表达、细胞增殖及对γ射线照射敏感性的影响。方法 以脂质体介导的HER2／neu基因转染MCF7细胞,用G418筛选阳性克隆。通过Western blot鉴定HER2／neu蛋白的表达,并检测p53、信号转导分子Akt和p-Akt蛋白含量的变化及PI3K通路抑制剂LY294002对上述蛋白表达水平的影响。以MTT法检测细胞增殖以及细胞对γ射线照射的敏感性。结果 共获得18个稳定转染HER2／neu基因的阳性克隆,其中1个克隆有HER2／neu基因过表达。过表达HER2／neu的MCF7细胞p-Akt蛋白含量升高,p53蛋白含量低于对照组细胞,LY294002能够抑制p-Akt蛋白和p53蛋白的变化。同时,过表达HER2／neu基因的MCF7细胞生长速度高于对照组细胞,对γ射线照射治疗的敏感性降低,而LY294002能够抑制细胞生长并增强放射治疗的敏感性。结论 MCF7细胞中HER2／neu基因的过表达,能够通过激活PI3K通路导致野生型p53蛋白含量减少、细胞增殖加快及放疗的敏感性降低,这可能是某些p53蛋白为野生型的肿瘤患者对治疗产生抗性的原因。     

Inhibition of activated phosphatidylinositol 3-kinase/AKT pathway in malignant pleural mesothelioma leads to G1 cell cycle arrest

Phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway plays pivotal roles in fundamental cellular functions including cell proliferation and cell survival. Its deregulation has been implicated in many types of human malignancies. We investigated the role of PI3K/AKT signaling pathway in human malignant pleural mesothelioma (MM). Here, we report that aberrant activation of the PI3K/AKT signaling pathway is associated with cell cycle progression in MM cells. Inhibition of the PI3K activity by its small molecule inhibitor LY294002 led to significant G1 cell cycle arrest and suppression of cell proliferation in all MM cell lines that we examined. In addition, we found that the protein level of p27Kip1 was up-regulated and the protein level of cyclin D1 was down-regulated following LY294002 treatment in those MM cell lines. However, no noticeable apoptosis induction was observed following 24 h of LY294002 treatment in those MM cell lines. These results confirm that the PI3K/AKT signaling pathway is aberrantly active and plays a critical role for the cell cycle progression in human MM cells.     

Combined Inhibition of PI3K and mTOR Exerts Synergistic Antiproliferative Effect,but Diminishes Differentiative Properties of Rapamycin in Acute Myeloid Leukemia Cells

A novel strategy has been suggested to enhance rapamycin-based cancer therapy through combining mammalian target of rapamycin (mTOR)-inhibitors with an inhibitor of the phosphatydilinositol 3-kinase PI3K/Akt or mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway. However, recent study demonstrated the potentiating effect of rapamycin on all-trans-retinoic acid (ATRA)-mediated differentiation of acute myelogenous leukemia (AML) cells, prompting us to investigate the effects of longitudinal inhibition of PI3K/Akt/mTOR signaling pathway on both proliferation and differentiative capacity of AML. In NB4, HL-60, U937 and K562 cell lines, rapamycin exerted minimal antiproliferative effects, and combining PI3K inhibitor LY 294002 and rapamycin inhibited proliferation more than LY 294002 alone. Rapamycin potentiated differentiation of ATRA-treated NB4 cells, but the combination of rapamycin and LY 294002 inhibited the expression of CD11b in both ATRA- and phorbol myristate acetate (PMA)-stimulated cells more than PI3K inhibitor alone. These results demonstrate that, although the combination of PI3K inhibitor and rapamycin is more effective in inhibiting proliferation of AML, the concomitant inhibition of PI3K and mTOR by LY 294002 and rapamycin has more inhibitory effects on ATRA-mediated differentiation than the presence of PI3K-inhibitor alone, and diminishes positive effects of rapamycin on leukemia cell differentiation.     

低表达DJ-1调控PI3K/AKT通路对肺癌细胞增殖、凋亡的影响

目的:探讨低表达DJ-1对肺癌细胞增殖凋亡及PI3K/AKT通路的影响。方法:采用Western blot检测肺癌细胞系中DJ-1蛋白的表达;以脂质体法转染干扰SK-MES-1细胞中DJ-1蛋白的表达后,MTT法检测细胞的增殖变化,流式细胞仪检测细胞的凋亡情况,Western blot检测细胞中p-AKT和AKT蛋白的表达水平;将PI3K/AKT通路抑制剂LY294002处理SK-MES-1细胞48 h后,MTT法和流式细胞仪分别检测细胞的增殖和凋亡情况,Western blot检测细胞中p-AKT和AKT蛋白的表达。结果:与正常肺上皮BEAS-2B细胞相比,肺腺癌A549细胞和肺鳞癌SK-MES-1细胞中DJ-1蛋白的相对表达量均显著升高,且SK-MES-1细胞中DJ-1蛋白的表达量高于A549细胞,差异均有统计学意义（P＜0.05）。转染后siRNA DJ-1组细胞中DJ-1蛋白的表达量明显低于对照组（P＜0.05）;与对照组相比,转染24 h后siRNA DJ-1组细胞的吸光值（OD值）变化不显著（P＞0.05）,而转染48 h和72 h后细胞的OD值明显降低（P＜0.05）;转染48 h后,与对照组相比,siRNA DJ-1组细胞的凋亡率显著升高（P＜0.05）,p-AKT/AKT值显著降低（P＜0.05）。SK-MES-1细胞经抑制剂LY294002处理48 h后,细胞的增殖凋亡趋势与下调DJ-1表达的结果相一致。结论:DJ-1在肺癌细胞中高表达,下调其表达能够抑制细胞增殖,促进细胞凋亡,其作用机制可能与PI3K/AKT信号通路有关。     

NOTCH通路依赖PI3K/AKT通路调控胶质瘤干细胞增殖和自我更新能力

目的 探讨NOTCH通路对胶质母细胞瘤细胞增殖和自我更新能力调控及其潜在机制。方法 体外培养LN-229、U118-MG和A172胶质瘤干细胞球;MTT实验和单细胞成球实验分别检测胶质瘤干细胞增殖和自我更新能力;靶向AKT1慢病毒载体shRNA和PI3K抑制剂LY294002抑制PI3K/AKT信号通路活性,检测NOTCH通路功能是否依赖PI3K/AKT通路;Western blot检测NOTCH通路和PI3K/AKT通路活性;BLISS独立模型评价NOTCH抑制剂DAPT和LY294002的相互作用。结果 LN-229、U118-MG和A172体外培养形成胶质瘤干细胞球;DAPT抑制NOTCH通路活性同时降低PI3K/AKT通路活性,且NOTCH通路对PI3K/AKT通路的调控不依赖于PTEN介导;DAPT抑制胶质瘤干细胞增殖和体外单细胞成球能力;ShAKT1和LY294002抑制PI3K/AKT通路活性,同时显著减弱DAPT对胶质瘤干细胞增殖和体外单细胞成球能力的抑制作用（P<0.05）,DAPT和LY294002联合使用产生拮抗作用。结论 NOTCH通路依赖于PI3K/AKT通路调控胶质瘤干细胞增殖和自我更新能力。     

LY294002对LK2和H460肺肿瘤细胞系细胞增殖及SKP2和p27表达的影响

目的:探讨PI3K抑制剂LY294002对肺肿瘤细胞增殖影响及其分子机制。方法:培养肺癌细胞系LK2和H460,LY294002阻断PI3K信号通路后,MTT比色法测定细胞生长曲线;FCM分析细胞周期变化;实时RT-PCR、蛋白印迹法分别观察PI3K信号下游蛋白SKP2、p27mRNA及蛋白表达变化。结果:与对照组相比,处理组细胞生长速率受抑制,细胞周期阻滞于G1期。实时RT-PCR及蛋白印迹结果显示,LY294002处理后SKP2mRNA、蛋白表达水平均降低,2种细胞中基因降低量分别为42%和41%,蛋白降低量为65%和55%,p27mRNA表达水平增加了49%和36%,但蛋白表达水平不变。结论:肺肿瘤细胞LK2和H460中PI3K通路通过SKP2,但并不经由p27调节细胞周期,调控细胞增殖。     

Cross‐signaling among phosphinositide‐3 kinase,mitogen‐activated protein kinase and sonic hedgehog pathways exists in esophageal cancer

The hedgehog (Hh) signaling pathway is essential for the development of tissues and organs. Hyperactive Hh signaling has been implicated in many gastric cancers, including esophageal cancer. However, the interaction between the Hh pathway and other potential signaling pathways in primary esophageal tumorigenesis has not been well investigated. In our study, we found that esophageal cancer cells expressed Hh signaling molecules and that the hyperexpression of Hh target genes was related to protein kinase B (AKT) activation but not extracellular signal‐regulated kinase activation. We analyzed the relationship between Gli1 or p‐AKT expression and clinicopathological features in esophageal carcinoma samples and found that Gli1 expression was associated with lymph vessel invasion (p = 0.016), blood vessel invasion (p = 0.006) and a poor prognosis (p = 0.003), and p‐AKT expression was associated with blood vessel invasion (p = 0.031) and a poor prognosis (p = 0.031). We also studied the relationship between Hh and phosphinositide‐3 kinase (PI3K)/AKT or mitogen‐activated protein kinase (MAPK) signaling pathways in both TE‐1 and TE‐10 cell lines. We found that the PI3K/AKT pathway played a critical role in Hh signaling after stimulation with epidermal growth factor, Gβγ and N‐Shh. Conversely, PI3K/AKT and MAPK signaling cooperated with the Shh pathway to promote esophageal cancer cell survival and proliferation. The results from esophageal cancer cells shed light on the significance of Hh signaling in esophageal tumor formation and the crosstalk of the Hh pathway with other basic signaling pathways, which is consistent with that observed in human tumor samples.     

AKT and mTOR phosphorylation is frequently detected in ovarian cancer and can be targeted to disrupt ovarian tumor cell growth

Activation of the PI3K/AKT pathway may contribute to tumorigenesis. AKT mediates survival signals that protect cells from apoptosis and, thus, is a potentially important therapeutic target. To determine the frequency of AKT activation in human ovarian cancer, we screened a tumor tissue microarray with a phospho-specific pan-AKT (Ser473) antibody, which revealed elevated staining in 21 of 31 (68%) ovarian carcinomas. Phospho-AKT staining was associated with that of phospho (active)-mTOR in 27 of 31 (87%) ovarian tumors, with 17 (55%) tumors showing elevated phospho-mTOR positivity. We tested the effects of AKT/mTOR activation on the therapeutic sensitivity of ovarian cancer cells. Pretreatment of SKOV3 cells, which exhibit constitutive AKT activity under low serum conditions, with the PI3K inhibitor LY294002 augmented cisplatin-induced apoptosis. In contrast, ovarian cancer cell lines OVCAR4 and OVCAR5, which have low basal levels of AKT activity, did not show increased cisplatin-induced apoptosis when pretreated with LY294002. In addition, inhibition of mTOR activity with rapamycin resulted in G1 arrest in SKOV3 cells, but not in OVCAR4 or OVCAR5 cells. Collectively, these findings indicate that active AKT and downstream mTOR represent potentially important therapeutic and/or chemopreventive targets in ovarian cancer.