Phosphatase And Tensin Homolog Deleted On Chromosome Ten Expression Research Articles

Use of antioxidant nanoliposomes for co-delivery of PTEN plasmids and plumbagin to induce apoptosis in hepatic cancer cells.

Hepatocellular carcinoma remains a challenging contributor to the global cancer and related mortality, and claims approximately 800,000 deaths each year. Dysregulation or loss of function mutations involving the tumor suppressor gene, phosphatase and tensin homolog deleted on chromosome ten (PTEN), has been well-characterized in various cancers to elicit anomalous cell proliferation and oncogenic transformation. However, the delivery and bioavailability of genes/drugs of interest to carcinomas remains a serious bottleneck behind the success of any anti-cancer formulation. In this study, we have engineered nanoliposomes containing PTEN plasmids, plumbagin, and antioxidant cerium oxide nanoparticles (Lipo-PTEN-Plum) to restore the PTEN expression and inhibit the AKT/PI3K pathway. The Lipo-PTEN-Plum was quasi-spherical in shape with ∼110 nm diameter and ∼64% plumbagin loading efficiency. The Lipo-PTEN-Plum was successfully internalized HepG2 cells, restore PTEN expression and inhibit PI3K/AKT pathway to induce death in cells grown in monolayer and in form of spheroids. Mechanistically, the formulation showed G2/M cell cycle arrest, DNA damage and apoptosis in hepatic cancer cells. Other cellular events such as Caspase-7 overexpression and PI3K (phosphoinositide 3-kinase), AKT (a serine/threonine protein kinase), PARP [Poly (ADP-ribose) polymerases], and mTOR (Mammalian target of rapamycin) inhibition led to the apoptosis in hepatic cancer cells. The mRNA expression profile of PTEN, PI3K, AKT3, Caspase-7, PARP and mTOR proteins, primarily controlling the cancer cell proliferation and apoptosis, suggest that exogenous supply of PTEN could regulate the expression of oncogenic proteins and thus cancer progression.

HBX suppresses PTEN to promote the malignant progression of hepatocellular carcinoma through mi-R155 activation

Introduction and objectivesHepatocellular carcinoma (HCC) is one of the most common and fatal tumors in the world, ranking third in cancer-related mortality. Chronic HBV infection is one of the major risk factors for hepatocellular carcinoma in China, Korea, and Sub-Saharan Africa. The HBx protein encoded by the X gene of HBV is a broadly regulated protein involved in transcriptional activation, epigenetics, apoptosis, DNA repair, and other regulatory processes. This study aimed to investigate the mechanism of HBx regulation of miR-155 and PTEN (Phosphatase and tensin homolog deleted on chromosome ten) in HBV-HCC. MethodsExosomal miR-155 quantity was analyzed by sampling serum exosomes of patients with hepatocellular carcinoma and normal subjects. The analysis was divided into different subgroups according to HBV positivity or negativity. At the cellular level, the biological roles of HBX, microRNA-155 and PTEN on hepatocellular carcinoma cells and their regulatory relationships with each other were verified. ResultsMicroRNA-155 and PTEN expression in HBV-positive HCC liver cancer tissues were negatively correlated, and HBX and miR-155 expression were positively correlated; microRNA-155 could target and inhibit PTEN expression, thereby promoting hepatocellular carcinoma cell activity, inhibiting apoptosis, and promoting invasion and migration; HBX could upregulate microRNA-155 thereby inhibit PTEN to promote malignant transformation of hepatocellular carcinoma. ConclusionsHBX could promote malignant transformation of hepatocellular carcinoma cells by upregulating microRNA-155 expression and thereby inhibiting the PTEN/PI3K-AKT pathway. Blocking miR-155 expression could attenuate the proliferation-promoting and invasive effects of HBX.

Percutaneous Intracoronary Delivery of Plasma Extracellular Vesicles Protects the Myocardium Against Ischemia-Reperfusion Injury in Canis.

[Figure: see text].

GAS5 promotes podocyte injury in sepsis by inhibiting PTEN expression.

To investigate the potential role of long noncoding RNA (lncRNA) growth arrest specific transcript 5 (GAS5) in sepsis-induced podocyte injury and its underlying mechanism. The sepsis model was established by lipopolysaccharide (LPS) induction in podocytes. The expression levels of Nephrin and GAS5 were detected by quantitative Real-time polymerase chain reaction (qRT-PCR) after LPS induction in podocytes for 12 h, 24 h and 36 h, respectively. Western blot was used to detect the expression level of Nephrin in sepsis-induced podocytes. The mRNA expressions of GAS5 and Nephrin in podocytes were detected after transfection of GAS5 siRNA. Albumin influx in podocytes after GAS5 knockdown was detected by Transwell assay. Western blot was used to detect the protein expression of Snail in sepsis after GAS5 knockdown. The target gene of GAS5 was predicted by bioinformatics analysis. QRT-PCR and Western blot were used to detect the protein and mRNA levels of PTEN (phosphatase and tensin homolog deleted on chromosome ten). Nephrin expression and the albumin inflow after PTEN knockdown were then measured. The expression of PI3K/AKT/GSK3β was also detected after GAS5 was downregulated while PTEN was upregulated. LPS stimulation downregulated the mRNA expression of Nephrin in podocytes and achieved the lowest level at 24 h. The protein expression change of Nephrin was consistent with its mRNA expression. In the septic state, the albumin influx of podocytes remarkably increased, but the function of podocyte barrier was weakened. Besides, GAS5 expression decreased in a time-dependent manner in LPS-induced podocytes. After GAS5 knockdown by siRNA, Nephrin expression and the function of podocyte barrier were significantly reduced. Snail expression was also upregulated in septic state, and GAS5 knockdown increased the expressions of phosphorylated Snail and PI3K/AKT/GSK3β. After knockdown of GAS5, the mRNA and protein levels of PTEN significantly decreased, which was contract to the expression of Snail. However, overexpression of PTEN could reverse the promotive effect of GAS5 on PI3K/AKT activation. GAS5 expression decreased in sepsis-induced podocyte injury, and GAS5 was involved in regulating sepsis-induced podocyte injury by reducing PTEN expression.

Triptolide impairs thioredoxin system by suppressing Notch1-mediated PTEN/Akt/Txnip signaling in hepatocytes

Triptolide (TP) is the main ingredient of Chinese herb Tripterygium wilfordii Hook f. (TWHF). Despite of its multifunction in pharmaceutics, accumulating evidences showed that TP caused obvious hepatotoxicity in clinic. The current study investigated the role of Notch1 signaling in TP-induced hepatotoxicity. Our data indicated that TP inhibited the protein expression of Notch1 and its active form Notch intracellular domain (NICD) leading to increased PTEN (phosphatase and tensin homolog deleted on chromosome ten) expression. Moreover, PTEN triggered Txnip (thioredoxin-interacting protein) activation by inhibiting Akt phosphorylation, which resulted in reduction of Trx (thioredoxin). In conclusion, TP caused liver injury through initiating oxidative stress in hepatocyte. This study indicated the potency of Notch1 to protect against TP-induced hepatotoxicity.

MicroRNA-200a promotes proliferation and invasion of ovarian cancer cells by targeting PTEN.

We investigate whether microRNA-200a could regulate proliferation and invasion of ovarian cancer cells, thereby participating in the occurrence and development of ovarian cancer. We also explore the specific mechanism of microRNA-200a in regulating ovarian cancer. Expression level of microRNA-200a in ovarian cancer tissues and paracancerous tissues were detected by quantitative Real-time polymerase chain reaction (qRT-PCR). The regulatory effects of microRNA-200a on proliferation and invasion of ovarian cancer cells were examined by Cell counting kit-8 (CCK-8) and cell invasion assay, respectively. Dual-luciferase reporter gene assay was performed to confirm the binding relationship between microRNA-200a and PTEN (phosphatase and tensin homolog deleted on chromosome ten). The regulatory role of microRNA-200a in PTEN expression was accessed by Western blot. Rescue experiments were conducted to assess whether microRNA-200a regulated proliferation and invasion of ovarian cancer cells by inhibiting PTEN expression. MicroRNA-200a expression in ovarian cancer tissues was significantly higher than that of paracancerous tissues. Besides, microRNA-200a was also overexpressed in ovarian cancer cell lines than that of normal ovarian cells. Overexpression of microRNA-200a promoted the proliferative and invasive abilities of SKOV3 and OVCAR3 cells. Dual-luciferase reporter gene assay showed that microRNA-200a could directly degrade PTEN. Overexpression of PTEN in SKOV3 and OVCAR3 cells partially reversed the increased cell proliferation and invasion induced by overexpressed microRNA-200a. Overexpressed microRNA-200a promoted the proliferative and invasive abilities of ovarian cancer cells, which might be related to the targeted regulation of PTEN expression.

The expression and significance of Tipα and PTEN in gastric cancer

Objective To investigate the correlation between tumor necrosis factor -α inducing protein (Tipα), phosphatase and tensin homologue deleted onchromosome ten (PTEN) and gastric cancer. Methods To detecte the relative quantification of Tipα and lost phosphatase gene of tenth chromosome PTEN in gastric cancer, paracancerous tissues and normal gastric tissues by real-time quantitative polymerase chain reaction (RT-qPCR) technique. Results ⑴ The expression levels of Tipα and PTEN in gastric cancer, paracancerous tissues and normal gastric tissues were 0.83±0.07, 0.16±0.10, 0.10±0.12, 0.23±0.05, 0.92±0.07, 1.84±0.13, respectively. Comparison of gastric cancer with paracancerous tissues and normal gastric tissue, there were significant statistical differences(P<0.05). ⑵ The expression levels of Tipα and PTEN in low and well differentiated gastric carcinoma were1.10±0.04, 0.36±0.05, 0.08±0.05, 0.36±0.04, respectively, both of which are significantly associated with the degree of differentiation of gastric cancer(P<0.05); The expression levels of Tipα and PTEN in gastric cancer tissues with or without lymph node metastasis were 0.18±0.12, 0.82±0.09, 0.10±0.12, 0.38±0.10, both had significant correlation with lymph node metastasis of gastric cancer(P<0.05); The expression of PTEN in gastric cancer tumor node metastasis (TNM)Ⅰ-Ⅱ and Ⅲ-Ⅳ were 0.25±0.04, 0.06±0.07, which means PTEN was significantly correlated with gastric cancer TNM staging(P<0.05). ⑶ The expressions of Tipα and PTEN were negatively correlated in gastric cancer tissues (r=-0.883, P<0.05). Conclusions ⑴ Tipα may be a cancer-promoting factor; PTEN maybe a tumor suppressor factor; ⑵ There is a negative correlation between the expression of Tipα and PTEN in gastric carcinoma. Key words: Stomach neoplasms/PA/ME; Tumor necrosis factor-alpha-inducing protein/ME; PTEN phosphohydrolase/ME

PI3K/Akt/mTOR signaling &amp; its regulator tumour suppressor genes PTEN &amp; LKB1 in human uterine leiomyomas

Background & objectives:Despite their high occurrence and associated significant level of morbidity manifesting as spectrum of clinical symptoms, the pathogenesis of uterine leiomyomas (ULs) remains unclear. We investigated expression profile of tumour suppressor genes PTEN (phosphatase and tensin homolog deleted on chromosome ten) and LKB1 (liver kinase B1), and key signaling components of P13K (phosphatidylinositol 3-kinase)/Akt (protein kinase B)/mTOR (mammalian target of rapamycin) pathway in leiomyomas and adjacent normal myometrium in women of reproductive age, to explore the possibility of targeting this pathway for future therapeutic implications.Methods:Real time PCR (qPCR) was used to quantify relative gene expression levels of PTEN, Akt1, Akt2, mTOR, LKB1 and VEGFA (vascular endothelial growth factor A) in leiomyoma as compared to adjacent normal myometrium. Immunohistochemistry was subsequently performed to analyze expression of PTEN, phospho-Akt, phospho-mTOR, phospho-S6, LKB1 and VEGFA in leiomyoma and adjacent normal myometrium.Results:Significant upregulation of PTEN (2.52 fold; P=0.03) and LKB1 (3.93 fold; P=0.01), and downregulation of VEGFA (2.95 fold; P=0.01) genes were observed in leiomyoma as compared to normal myometrium. Transcript levels of Akt1, Akt2 and mTOR did not vary significantly between leiomyoma and myometrium. An increased immunoexpression of PTEN (P=0.015) and LKB1 (P<0.001) and decreased expression of VEGFA (P=0.01) was observed in leiomyoma as compared to myometrium. Immunostaining for activated (phosphorylated) Akt, mTOR and S6 was absent or low in majority of leiomyoma and myometrium.Interpretation & conclusions:Upregulation of PTEN and LKB1 in concert with negative or low levels of activated Akt, mTOR and S6 indicates that PI3K/Akt/mTOR pathway may not play a significant role in pathogenesis of leiomyoma.

Expression and clinical evidence of miR-494 and PTEN in non-small cell lung cancer.

The aim of this study was to explore the expression and clinical significance of miR-494 and PTEN (phosphatase and tensin homologue deleted on chromosome ten) in non-small cell lung cancer (NSCLC). Immunohistochemistry for PTEN and in situ hybridization (ISH) for miR-494 were performed in 92 NSCLC tissues and 10 normal lung tissues to detect their expression, and correlation between their expression with clinical characteristics and prognosis was analyzed. The expression of miR-494 was significantly higher in NSCLC than in normal lung tissues (P = 0.004). The positive expression of PTEN protein in the lung carcinoma tissues was significantly lower than that in the normal lung tissues (P = 0.013), while the level of miR-494 expression was negatively correlated with PTEN expression (r = -0.577, P < 0.01). The high positive rate of miR-494 was positively correlated with pathological TNM (p-TNM) staging and lymph node metastasis. The expression of miR-494 was negatively correlated with grade of differentiation. However, the expression of PTEN was positively correlated with grade of differentiation. Patients with over-expression of miR-494 had a shorter overall survival (OS), while the negative group of PTEN was correlated with poor OS. MiR-494 over-expression and low PTEN expression are closely related to tumor p-TNM staging and lymph node metastasis, differentiation, and OS. Combined detection of PTEN and miR-494 can aid in determining malignancy degree and the prognosis of patients with NSCLC. MiR-494 may be served as a novel prognostic factor and may lead to new treatment strategies for NSCLC.

ASSA14-03-02 Protective Effects of Paeoniflorin against Myocardial Fibrosis in Isoprenaline-induced Chronic Heart Failure Rats

<h3>Background</h3> The life time risk of developing heart failure is 20% for Americans over 40 years old. Chronic heart failure (CHF) is becoming a global health problem in the elderly. Previous studies indicate that immune-inflammatory activation plays an important role in the pathogenesis of myocardial fibrosis in CHF. Paeoniflorin (PF) is one of the main effective components of peony, which has been widely used in traditional Chinese medicine for its potent immunomodulatory and anti-inflammatory properties. However, the effects of PF on myocardial fibrosis remain unclear. <h3>Objective</h3> To exam whether PF protects myocardial fibrosis and improves cardiac function in isoprenaline-induced CHF rats. <h3>Methods</h3> Age matched male Wistar rats were used in this study. CHF and cardiac remodelling model was established by hypodermic injection of isoprenaline for ten days. After the CHF model establishment, the rats were randomly divided to receive placebo, PF (50 or 100 mg/kg/d, i.g.) or captopril for six weeks. Cardiac remodelling markers including ventricular mass, collagen volume fraction, perivascular collagen area and hydroxyproline concentration were studied. Echocardiography, Masson staining, immunohistochemistry, western blot and real-time PCR were performed. The protein and mRNA expression of PTEN (phosphatase and tensin homolog deleted on chromosome ten) were analysed. <h3>Results</h3> Compared to the untreated CHF rats, the perivascular collagen area, collagen volume fraction (30.97 ± 4.22% vs 13.75 ± 3.77%), hydroxyproline concentration (4.92 ± 0.78 μg/mg wet weight vs 2.08 ± 0.43 μg/mg wet weight), LVIDd (8.40 ± 0.52 mm vs 6.11 ± 0.58 mm) and LVIDs (6.44 ± 0.93 mm vs 4.49 ± 0.38 mm) were significantly lower in PF-treated (100 mg/kg/d) rats (p &lt; 0.05, n = 8). PF treatment also improved LVEF levels (46.44 ± 3.50% vs 60.89 ± 3.10%). Besides, the expressions of PTEN protein (0.11 ± 0.05 vs 0.63 ± 0.19, p &lt; 0.05) and mRNA (0.22 ± 0.06 vs 0.39 ± 0.02, p &lt; 0.01) were higher in PF-treated rats (n = 3). No damage of renal or liver function was detected in these rats (p &gt; 0.05). Furthermore, we also found that PF can accelerate myocardial fibroblast cell apoptosis and upregulate PTEN signalling <i><i>in vitro</i></i>. <h3>Conclusions</h3> Our findings indicate that PF could alleviate myocardial fibrosis and improve cardiac function in isoprenaline-induced CHF rats by upregulating PTEN signalling pathway. PF may be a potential therapeutic drug for CHF.

ASSA14-03-43 Novel therapeutic uses of triptolide in reversing isoprenaline-induced cardiac remodelling in Wistar rats

Background Nowadays, chronic heart failure (CHF) is a global health problem in the elderly. Over 650 thousand new cases are diagnosed every year in the United States. Previous studies indicate that immune-inflammatory activation plays an important role in the pathogenesis of cardiac remodelling in CHF patients. PG-490, triptolide (TPL), is a diterpene triepoxide with potent immunosuppressive and anti-inflammatory properties and has been used in China for centuries to treat immune-related disorders. However, the effects of TPL on CHF and cardiac remodelling remain unclear. Objective To determine whether TPL can protect myocardial fibrosis and improve cardiac function in isoprenaline-induced CHF rats. Methods Age matched male Wistar rats were used in this study. CHF and cardiac remodelling model was established by hypodermic injection of isoprenaline for ten days. The rats were randomly divided to receive placebo, TPL (20 or 100 μg/kg/d, i.g.) or captopril for six weeks. Cardiac remodelling markers including ventricular mass, collagen volume fraction, perivascular collagen area, collagenI and hydroxyproline concentration were studied. Echocardiography, Masson staining, immunohistochemistry, western blot and real-time PCR were performed. The protein and mRNA expression of PTEN (phosphatase and tensin homolog deleted on chromosome ten) were analysed. Results Compared to the untreated CHF rats, the cardiac remodelling markers, LVEDD (8.40 ± 0.52 mm vs 6.11 ± 0.58 mm) and LVESD (6.44 ± 0.93 mm vs 4.49 ± 0.38 mm) were significantly lower in TPL-treated (100 μg/kg/d) rats (p Figure 1 ) and mRNA (0.22 ± 0.06 vs 0.52 ± 0.01) were higher in TPL-treated rats (p in vitro . Conclusions Our findings indicate that TPL could alleviate myocardial remodelling and improve cardiac function in rats by upregulating PTEN signalling pathway. TPL may be a potential therapeutic drug for CHF.

MiR-205 inhibits cell apoptosis by targeting phosphatase and tensin homolog deleted on chromosome ten in endometrial cancer Ishikawa cells.

BackgroundMicroRNAs (miRNAs) are frequently dysregulated in human cancers and can act as either potent oncogenes or tumor suppressor genes. In the present study, we intend to prove that the gene PTEN (phosphatase and tensin homolog deleted on chromosome ten) is a target gene of miR-205 and to investigate the suppressive effects on PTEN transcriptional activity by enhancing miR-205 expression in endometrial cancer Ishikawa cells.MethodsUsing Ishikawa cells as model systems, we up-regulated miR-205 expression by transient transfection with miR-205 mimics. A luciferase reporter assay, qRT-PCR and western blotting assays were used to verify whether PTEN is a direct target of miR-205. Meanwhile, the modulatory role of miR-205 in the AKT (protein kinase B) pathway was evaluated by determining the AKT phosphorylation. As a biological counterpart, we investigated cell apoptosis using flow cytometry.ResultsOur data indicate that miR-205 down-regulates the expression of PTEN through direct interaction with the putative binding site in the 3′-untranslated region (3′-UTR) of PTEN. Moreover, we documented the functional interactions of miR-205 and PTEN, which have a downstream effect on the regulation of the AKT pathway, explaining, at least in part, the inhibitory effects on Ishikawa cell apoptosis of enhancing miR-205 expression.ConclusionsFor the first time, we demonstrate that the expression of PTEN is directly regulated by miR-205 in endometrial cancer cells and leads the inhibition of cellular apoptosis. This relationship could be targeted for new therapeutic strategies for endometrial cancer.

Hypertrophied Myocardium Is Refractory to Sevoflurane-Induced Protection With Alteration of Reperfusion Injury Salvage Kinase/Glycogen Synthase Kinase 3β Signals

Recent studies have demonstrated that volatile anesthetic postconditioning confers myocardial protection against ischemia-reperfusion injury through activation of the reperfusion injury salvage kinase (RISK) pathway. As RISK has been shown to be impaired by ventricular hypertrophy, we investigate whether anesthetic-induced cardiac protection was maintained in rat hearts with ventricular hypertrophy. Transverse aortic constriction operation was performed on male Sprague-Dawley rats to induce left ventricular (LV) hypertrophy, then sham-operated or hypertrophied rat hearts were subjected to 40 min of global ischemia and 2 h of reperfusion. The isolated hearts received 3% sevoflurane for 10 min or six cycles of 10-s ischemia/10-s reperfusion after reperfusion. The hemodynamics, infarct size, PTEN (phosphatase and tensin homolog deleted on chromosome ten), phosphorylated Akt, phosphorylated extracellular regulated protein kinase (ERK) 1/2, and phosphorylated glycogen synthase kinase 3β (GSK3β) were determined. We found the myocardial expression of PTEN, phosphorylated Akt, ERK1/2, and phosphorylated GSK3β did not significantly differ between sham-operated and transverse aortic constriction-control groups. Both sevoflurane and ischemic postconditioning significantly improved LV hemodynamics, reduced infarct size, and increased the phosphorylation of Akt, ERK1/2, and their downstream target of GSK3β in the sham-operated rat hearts. In contrast, neither sevoflurane nor ischemic postconditioning improved LV hemodynamic, reduced infarct size, and increased the phosphorylated Akt, ERK1/2, and GSK3β in hypertrophied myocardium. All the results above indicate that ventricular hypertrophy abrogated sevoflurane-induced cardioprotection against ischemia-reperfusion injury by alteration of RISK/GSK3β signals.

Disruption of E3 ligase NEDD4 in peripheral neurons interrupts axon outgrowth: Linkage to PTEN

Exploiting molecules and pathways important in developmental axon behaviour may offer new insights into regenerative behaviour of adult peripheral neurons after injury. In previous work, we have provided evidence that inhibition or knockdown of PTEN (phosphatase and tensin homolog deleted on chromosome ten) dramatically increases adult peripheral axon outgrowth, especially in preconditioned neurons (Christie et al., 2010). PTEN appears to operate as an endogenous brake to regeneration. Recent reports from Drinjakovic et al. (2010) have highlighted a role for the ubiquitin proteasome system (UPS) during neurite outgrowth in developing Xenopus retinal ganglion cells. Specifically, disruption of the UPS E3 ligase Nedd4 (neural precursor cell-expressed developmentally down-regulated protein 4) inhibited neurite branching through up-regulation of PTEN. We explored the potential role of Nedd4 in the peripheral neurons of adult rat dorsal root ganglia (DRG), particularly its impact on regenerative behaviour. Global inhibition of the UPS in vitro was associated with a severe decrease in neurite branching, both in preconditioned (injured) and control DRG sensory neurons. These involved neurons however maintained or qualitatively increased their PTEN expression, suggesting ongoing PTEN activity during UPS inhibition. Considering component's of UPS more specifically, Nedd4 co-localized with PTEN within sensory neurons in vivo and in vitro. Nedd4 also co-localized with PTEN and NF200 labelled regenerating axons at the injury site in the periphery following a 3 day sciatic nerve cut. A significant role for this unique co-expression was observed with fluorescently tagged siRNA inhibition of Nedd4, which decreased neurite outgrowth, an impact associated with greater expression of PTEN and that was completely reversed with application of a PTEN inhibitor. Overall, our results suggest an important role for Nedd4 regulation of PTEN in the response of peripheral neurons to injury. By degrading PTEN among other potential actions, Nedd4 supports axonal outgrowth whereas its inhibition facilitates PTEN inhibition of regeneration.

Clinical significance of PTEN and B7-H1 expressions in pancreatic carcinoma

Objective To investigate the probable correlation between the expressions of phosphatase and tensin homologue deleted on chromosometen (PTEN) and B7-H1 protein in pancreatic carcinoma and the biological behavior characteristics of tumors. Methods Forty-three patients were recruited who had undergone surgical resection for pancreatic carcinoma between 2002 and 2009. The PTEN and B7-H1 protein expressions in the tissue specimens of these 43 patients and 5 non-pancreatic carcinoma people' s pancreatic tissue specimens were evaluated by immunohistochemistry ELPS technique, and the clinical and pathological features of these specimens and the follow-up information were analyzed. Results PTEN expressions were significantly lower in pancreatic carcinoma tissues than in non-pancreatic carcinoma people' s pancreatic tissues but B7-H1 expressions were significantly higher ( P ＜ 0. 01 ). The expression of PTEN was negatively correlated to that of B7-H1 (r = -0.414 ,P ＜0. 01). PTEN and B7-H1 expressions correlated with the pathological grade and tumor-node-metastasis ( TNM ) stage, peripancreatic invasion, regional lymph node involvement,respectively (P＜0. 05). B7-H1 expressions also significantly correlated with the ages (P＜0. 01). Furthermore, PTEN and B7-H1 expressions showed significant prognostic effects (P＜0.01) and there are correlations existed between combined PTEN/B7-H1 expression and prognostic effects (P ＜0. 05). Conclusion The expression of PTEN and B7-H1 may be significantly correlated to the carcinogenesis,development and prognosis of pancreatic carcinoma. Key words: Pancreatic cancer; PTEN; B7-H1; Biological behavior characteristics

Are we missing the mTOR target in breast cancer?

Abnormalities in the PI3K/Akt/mTOR intracellular signalling pathway have been implicated in several forms of human cancer. The central component of the pathway is the phosphatidylinositol 3-kinase (PI3K) heterodimer which comprises the p85 regulatory and p110 catalytic subunit, and mutations in the catalytic domain of PI3K have been identified in 20–25% of breast cancers [1, 2]. A further 15–35% of breast cancer patients demonstrate reduced expression of PTEN (phosphatase and tensin homologue deleted on chromosome ten), an endogenous inhibitor of the PI3K/AKT pathway [3]. Akt is the ultimate effector of the pathway and directs a multitude of intracellular effects related to growth, glucose metabolism, protein synthesis and cell survival. Activation of the PI3K/Akt pathway provides a cell with unrestricted growth and survival signals, and one important downstream consequence is the alleviation of the suppression by the tuberous sclerosis protein complex (TSC1/2) of the mammalian target of rapamycin (mTOR) [4]. There are at least two groups of mTOR proteins, and the key protein involved in transmitting signals from PI3K/Akt is the mTORC1 complex. Within cells, mTORC1 plays a critical role in the transduction of proliferative signals through phosphorylation of the translational regulator 4E-BP1 (eukaryotic initiation factor 4E-binding protein) and the ribosomal protein p70 (the 70-kDa S6 kinase) that facilitate translation of mRNAs into proteins [5, 6]. In recent years our understanding of this pathway in breast cancer has been exploited by the development of a number of targeted anti-cancer therapeutics, with mTOR inhibitors being the most clinically developed to date. Rapamycin is the original inhibitor of mTOR that was found to be a potent fungicide with immunosuppressive properties, and therapeutics were developed for clinical use in transplantation to prevent graft rejection. Subsequently rapamycin was found to have anti-proliferative effects in a range of experimental tumours [7], and rapamycin analogues with a more favourable pharmacological profile were synthesised including CCI-779 (temsirolimus), RAD001 (everolimus) and AP23573 (ridaforolimus). In various breast cancer cell lines with constitutive activation of the PI3K/Akt pathway due to either HER2 amplification, ER activation or PTEN deletions temsirolimus showed specific sensitivity [8], whilst everolimus (an oral hydroxyethyl ether of rapamycin) demonstrated impressive anti-proliferative activity against a wide variety of tumour models in vivo [9]. In the clinic mTOR antagonists were found to be relatively well tolerated in early phase I/II trials [10, 11], and development programmes were established in various solid malignancies, in particular renal cell cancer and breast cancer. But has the development of mTOR antagonists for breast cancer moved ahead too fast before it was really appreciated how best to utilise these drugs as targeted therapeutics? As a single agent temsirolimus showed only modest activity in the advanced breast cancer setting [12], and subsequently both temsirolimus and everolimus were developed in combination with endocrine therapy following encouraging evidence for additive or synergistic effects from several pre-clinical models of endocrine resistant breast cancer [13, 14]. However, results from randomised clinical trials of mTOR antagonists in combination with This is an invited commentary to articles doi: 10.1007/s10549-010-0967-z, 10.1007/s10549-010-0986-9, 10.1007/s10549-010-1058-x.

Upregulation of PTEN in Glioma Cells by Cord Blood Mesenchymal Stem Cells Inhibits Migration via Downregulation of the PI3K/Akt Pathway

BackgroundPTEN (phosphatase and tensin homologue deleted on chromosome ten) is a tumor suppressor gene implicated in a wide variety of human cancers, including glioblastoma. PTEN is a major negative regulator of the PI3K/Akt signaling pathway. Most human gliomas show high levels of activated Akt, whereas less than half of these tumors carry PTEN mutations or homozygous deletions. The unique ability of mesenchymal stem cells to track down tumor cells makes them as potential therapeutic agents. Based on this capability, new therapeutic approaches have been developed using mesenchymal stem cells to cure glioblastoma. However, molecular mechanisms of interactions between glioma cells and stem cells are still unknown.Methodology/Principal FindingsIn order to study the mechanisms by which migration of glioma cells can be inhibited by the upregulation of the PTEN gene, we studied two glioma cell lines (SNB19 and U251) and two glioma xenograft cell lines (4910 and 5310) alone and in co-culture with human umbilical cord blood-derived mesenchymal stem cells (hUCBSC). Co-cultures of glioma cells showed increased expression of PTEN as evaluated by immunofluorescence and immunoblotting assays. Upregulation of PTEN gene is correlated with the downregulation of many genes including Akt, JUN, MAPK14, PDK2, PI3K, PTK2, RAS and RAF1 as revealed by cDNA microarray analysis. These results have been confirmed by reverse-transcription based PCR analysis of PTEN and Akt genes. Upregulation of PTEN resulted in the inhibition of migration capability of glioma cells under in vitro conditions. Also, wound healing capability of glioma cells was significantly inhibited in co-culture with hUCBSC. Under in vivo conditions, intracranial tumor growth was inhibited by hUCBSC in nude mice. Further, hUCBSC upregulated PTEN and decreased the levels of XIAP and Akt, which are responsible for the inhibition of tumor growth in the mouse brain.Conclusions/SignificanceOur studies indicated that upregulation of PTEN by hUCBSC in glioma cells and in the nude mice tumors downregulated Akt and PI3K signaling pathway molecules. This resulted in the inhibition of migration as well as wound healing property of the glioma cells. Taken together, our results suggest hUCBSC as a therapeutic agent in treating malignant gliomas.

Promoter Methylation and Phosphorylation Status of PTEN in Taiwanese Breast Cancer.

Abstract Background: The PTEN (phosphatase and tensin homolog deleted on chromosome Ten) gene is a novel candidate of tumor suppressor that plays an important role in cell cycle regulation and apoptosis by modulating protein kinase-B/AKT activity. In western breast cancer population, PTEN downregulation is frequently associated with high-grade tumor, distant metastases and poorer disease-free survival. It is also noted that the methylation of the PTEN promoter results in PTEN inactivation in a subset of breast cancers. Taiwan is the low-incidence area of breast cancer. However, breast cancer has been jumped to the first place in the incidence of female malignancies in the past 3 years. Hence, we aim to explore the potential role of PTEN in Taiwanese breast cancer.Material and Method: Firstly, we used tissue microarrays containing 192 specimens (PTEN) and 193 samples (p-PTEN) from the Department of Pathology, Kaohsiung Medical University to analyze the expression patterns of PTEN and p-PTEN (Ser380/Thr382/Thr383) protein by immunohistochemistry (anti-PTEN antibody, A2B1, and anti-p-PTEN R, Santa Cruz). Clinical data, such as stage, grade, recurrence, lymph-node metastasis, and patient's age, tumor-specific survival (in months) and others were included in our statistic analysis. The genomic DNA (1 μg) obtained from 318 fresh specimens containing 159 matched cancer and non-cancer tissues were treated with sodium bisulfite for 16 hours and purified using methylation-specific PCR (MSP) kit (Millipore). Separate primers were used for amplification of methylated and unmethylated promoter regions. Primers that amplified a region of the promoter specific to PTEN gene but not the PTEN pseudogene were used. Primers used to amplify the methylated PTEN promoter were 5'-GTATTTCGAGTAAAGGAAGAAGACG-3' and 5'-GATAAAAAACTACAACC CAACGAA-3'; those used to amplify the unmethylated promoter were 5'- TATTTTGAGTA AAGGAAGAAGATGA-3' and 5'-CAATAAAAAACTACAACCCAACAAA-3'.Results: Our results showed that an increased nuclear expression of PTEN was positively correlated with an increased tumor staging (p=0.010). Furthermore, an increased p-PTEN expression in the cytoplasm was inversely correlated with tumor staging (p=0.003), while the decreased expression of p-PTEN in the nucleus was significantly correlated with positive recurrence rate (p=0.009) and metastasis to lymph node (p=0.033). Notably, the expression level of p-PTEN was positively correlated with that of PTEN. We also observed that the methylation of PTEN promoter was increased in 70% of breast cancer tissues as compared with the matched non-cancer breast tissues. Moreover, the increased methylation of PTEN promoter was positively correlated with tumor grade (p=0.037).Discussion: Taken together, our results suggest that an altered PTEN expression through methylation or phosphorylation may have potential roles in Taiwanese breast cancer. The underlying mechanism requires further investigation. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4060.

Aberrant Hyperactivation of Akt and Mammalian Target of Rapamycin Complex 1 Signaling in Sporadic Chordomas

Chordomas are rare, malignant bone neoplasms in which the pathogenic mechanisms remain unknown. Interestingly, tuberous sclerosis complex (TSC) is the only syndrome in which the incidence of chordomas has been described. We previously reported the pathogenic role of the TSC genes in TSC-associated chordomas. In this study, we investigated whether aberrant TSC/mammalian target of rapamycin complex 1 (mTORC1) signaling pathway is associated with sporadic chordomas. We assessed the status of mTORC1 signaling in primary tumors/cell lines of sacral chordomas and further examined upstream of mTORC1 signaling, including the PTEN (phosphatase and tensin homologue deleted on chromosome ten) tumor suppressor. We also tested the efficacy of the mTOR inhibitor rapamycin on signaling and growth of chordoma cell lines. Sporadic sacral chordoma tumors and cell lines examined commonly displayed hyperactivated Akt and mTORC1 signaling. Strikingly, expression of PTEN, a negative regulator of mTORC1 signaling, was not detected or significantly reduced in chordoma-derived cell lines and primary tumors. Furthermore, rapamycin inhibited mTORC1 activation and suppressed proliferation of chordoma-derived cell line. Our results suggest that loss of PTEN as well as other genetic alterations that result in constitutive activation of Akt/mTORC1 signaling may contribute to the development of sporadic chordomas. More importantly, a combination of Akt and mTORC1 inhibition may provide clinical benefits to chordoma patients.

Anti-tumor effect of honokiol alone and in combination with other anti-cancer agents in breast cancer

Honokiol, an active component isolated and purified from Chinese traditional herb magnolia, was demonstrated to inhibit growth and induce apoptosis of different cancer cell lines such as human leukaemia, colon, and lung cancer cell lines; to attenuate the angiogenic activities of human endothelial cells in vitro; and to efficiently suppress the growth of angiosarcoma in nude mice. In this study, we have demonstrated that treatment of different human breast cancer cell lines with honokiol resulted in a time- and concentration-dependent growth inhibition in both estrogen receptor-positive and -negative breast cancer cell lines, as well as in drug-resistant breast cancer cell lines such as adriamycin-resistant and tamoxifen-resistant cell lines. The inhibition of growth was associated with a G1-phase cell cycle arrest and induction of caspase-dependent apoptosis. The effects of honokiol might be reversely related to the expression level of human epidermal growth receptor 2, (HER-2, also known as erbB2, c-erbB2) since knockdown of her-2 expression by siRNA significantly enhanced the sensitivity of the her-2 over-expressed BT-474 cells to the honokiol-induced apoptosis. Furthermore, inhibition of HER-2 signalling by specific human epidermal growth receptor 1/HER-2 (EGFR/HER-2) kinase inhibitor lapatinib synergistically enhanced the anti-cancer effects of honokiol in her-2 over-expressed breast cancer cells. Finally, we showed that honokiol was able to attenuate the PI3K/Akt/mTOR (Phosphoinositide 3-kinases/Akt/mammalian target of rapamycin) signalling by down-regulation of Akt phosphorylation and upregulation of PTEN (Phosphatase and Tensin homolog deleted on chromosome Ten) expression. Combination of honokiol with the mTOR inhibitor rapamycin presented synergistic effects on induction of apoptosis of breast cancer cells. In conclusion, honokiol, either alone or in combination with other therapeutics, could serve as a new, promising approach for breast cancer treatment.