Xianghong Yang

Gastric cancer exosomes promote tumour cell proliferation through PI3K/Akt and MAPK/ERK activation

BACKGROUND Exosomes are nanometer-sized vesicles that are released by normal and neoplastic cells. Previous studies have focused on the interaction between tumour-derived exosomes and the immune system, as a consequence of immune suppression or enhancement. However, the effects of tumour-derived exosomes on tumour cells themselves have not been well studied. AIMS To investigate the effects of gastric cancer exosomes on tumour cell proliferation and the possible mechanisms. METHODS By serial centrifugation and sucrose gradient ultracentrifugation, we isolated and purified the exosomes from gastric cancer SGC7901 cells, then viewed them by electron microscopy. Cell proliferation was measured by 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide assay. Protein expression was assayed by Western blotting. RESULTS SGC7901-cell-derived exosomes promoted the proliferation of SGC7901 and BGC823 cells. The increase in proliferation induced by exosomes was accompanied by activation of Akt and extracellular-regulated protein kinase, and phosphoinositide 3-kinase or extracellular-regulated protein kinase inhibitor partially reversed the proliferative effect of exosomes. Moreover, the exosome-induced increase in activity of Akt and extracellular-regulated protein kinase coincided with decreased expression of the Casitas B-lineage lymphoma family of ubiquitin ligases. CONCLUSION Gastric cancer exosomes promoted tumour cell proliferation, at least in part, by activation of PI3K/Akt and mitogen-activated protein kinase/extracellular-regulated protein kinase pathways. The decreased expression of Casitas B-lineage lymphoma proteins might have contributed to the activation of Akt and extracellular-regulated protein kinase.

Oxaliplatin enhances TRAIL-induced apoptosis in gastric cancer cells by CBL-regulated death receptor redistribution in lipid rafts

Tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL) is a member of the tumor necrosis factor family that selectively induces apoptosis in cancer cells. However, gastric cancer cells are insensitive to TRAIL. In the present study, we show that oxaliplatin enhanced TRAIL‐induced apoptosis of MGC803, BGC823, and SGC7901 cells. Oxaliplatin promoted death receptor 4 (DR4) and death receptor 5 (DR5) clustering into aggregated lipid rafts, while the cholesterol‐sequestering agent nystatin partially prevented lipid raft aggregation, DR4 and DR5 clustering, and reduced apoptosis. Furthermore, the expression of the casitas B‐lineage lymphoma (Cbl) family was downregulated by oxaliplatin. Transfection of c‐Cbl or Cbl‐b partially reversed oxaliplatin‐induced lipid raft aggregation. These results indicated that oxaliplatin enhanced TRAIL‐induced gastric cancer cell apoptosis at least partially through Cbl‐regulated death receptor redistribution in lipid rafts.

Reversal of P-glycoprotein-mediated multi-drug resistance by the E3 ubiquitin ligase Cbl-b in human gastric adenocarcinoma cells†

P‐glycoprotein (P‐gp)‐mediated multi‐drug resistance (MDR) is a major barrier to the effective chemotherapy of many cancers. Recent studies have shown that inhibition of the PI3K/Akt signalling pathway can reverse P‐gp‐mediated MDR. We investigated the expression of activated Akt (p‐Akt) in 124 human gastric carcinoma tissue samples. Ubiquitous p‐Akt expression was recorded in the majority (88/124). There was a significant correlation between p‐Akt expression and the expression of P‐gp. In the adriamycin‐resistant MDR gastric carcinoma cell line SGC7901/ADR, p‐Akt expression was increased in comparison with the parental cell line SGC7901. Treatment of SGC7901/ADR cells with the PI3K inhibitor LY294002 reduced the expression of both p‐Akt and P‐gp. To explore the role of ubiquitin ligase Cbl‐b in this regulatory pathway, SGC7901/ADR cells were transfected with a plasmid overexpressing wild‐type Cbl‐b. This down‐regulated the expression of both p‐Akt and P‐gp. Furthermore, resistance to chemotherapeutic drugs was partially reversed. These results demonstrate an important role for Cbl‐b in reversing P‐gp‐mediated gastric cancer MDR through suppression of the PI3K/Akt signalling pathway and the down‐regulation of P‐gp expression. Copyright

The role of cbl family of ubiquitin ligases in gastric cancer exosome-induced apoptosis of Jurkat T cells.

Background. Exosomes are nanometer-sized vesicles with immunomodulatory functions, which are released by a diverse range of living cells. Although recent studies have shown that tumor-derived exosomes can suppress the function of T cells, the molecular mechanisms are not well understood. In the present study, we investigated the role of the Casitas B lineage lymphoma (cbl) family of ubiquitin ligases in gastric cancer exosome-induced apoptosis of Jurkat T cells. Materials and methods. By serial centrifugation and sucrose gradient ultracentrifugation, we isolated and purified the exosomes from gastric cancer SGC7901 cells, and identified them by electron microscopy and Western blotting. Cell apoptosis was detected using propidium iodide staining. Western blotting and RT-PCR was exploited to evaluate the expression of proteins and mRNA, respectively. Results. Gastric cancer exosomes induced Jurkat T cell apoptosis in a time- and dose-dependent manner and activated caspases 3, 8 and 9. The expression of Cbl-b and c-Cbl was up-regulated during exosome-induced apoptosis of cells. Meanwhile, exosomes induced ubiquitination of the p85 subunit of phosphoinositide 3-kinase (PI3K) and reduced downstream Akt activity. Inhibition of proteasome led to partial restoration of Akt activity and cell apoptosis. Discussion and Conclusions. The Cbl family of ubiquitin ligases might be involved in regulation of exosome-induced apoptosis of Jurkat T cells by increasing PI3K proteasome degradation, inactivation of PI3K/Akt signaling, thus mediating some effects of caspase activation.

Interferon-α sensitizes human gastric cancer cells to TRAIL-induced apoptosis via activation of the c-CBL-dependent MAPK/ERK pathway

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the tumor necrosis factor family that induces apoptosis in cancer cells. However, gastric cancer cells are insensitive to TRAIL. In the present study, we show that pretreatment with IFN-α enhanced TRAIL-induced apoptosis of gastric cancer MGC803 cells. IFN-α up-regulated death receptor 5 (DR5) expression and down-regulated survivin expression. Furthermore, extracellular-regulated protein kinase (ERK1/2) activation was induced by IFN-α, and a combination of IFN-α and TRAIL led to further activation of ERK1/2. Inhibition of the MAPK/ERK signaling pathway partially reversed apoptosis, as well as the expression patterns of DR5 and survivin. Moreover, the expression of the c-casitas B-lineage lymphoma (c-Cbl) family was down-regulated by IFN-α. Transfection of c-Cbl suppressed IFN-α-induced ERK activation. These results indicate that IFN-α enhances TRAIL-induced apoptosis in gastric cancer cells at least partially via downregulation of c-Cbl, and subsequent up-regulation of the MAPK/ERK pathway.

Receptor activator for nuclear factor κ B expression predicts poor prognosis in breast cancer patients with bone metastasis but not in patients with visceral metastasis

Background It was recently reported that receptor activator for nuclear factor κ B ligand (RANKL)/receptor activator for nuclear factor κ B (RANK) pathway is critical for RANK-expressing cancer cells to home to bone and associates with disease progression of cancer. The present study was aimed to evaluate the effect of RANK on prognosis in breast cancer patients with bone metastasis and patients with visceral metastasis. Methods Immunohistochemical staining for RANK was carried out on paraffin-embedded primary tumour tissue sections from 102 patients with metastatic breast cancer. Survival analysis and Cox proportional hazards model were used to explore the prognostic value of RANK expression in breast cancer. Results The RANK expression rates were 47.1% in metastatic breast cancer. Patients with RANK expression showed significantly poor progression-free survival and disease-specific survival. Subgroup analysis demonstrated that the significant difference in prognosis completely resulted from the occurrence of bone metastasis. Multivariate analysis demonstrated that RANK expression was an independent predictor of bone metastasis-free survival and disease-specific survival in patients with bone metastasis. Conclusions RANK expression might be an independent predictor of poor prognosis in breast cancer patients with bone metastasis, and RANK expression does not associate with the prognosis in patients with visceral metastasis.

Elevated serum CA19-9 level is a promising predictor for poor prognosis in patients with resectable pancreatic ductal adenocarcinoma: a pilot study

BackgroundPancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive human cancers. Several studies have reported that the carbohydrate antigen 19-9 (CA19-9) level is a useful marker for predicting the prognosis for PDAC after resection. However, the cutoff value of CA19-9 used to predict prognosis varied among these reports. The aims of this study were to evaluate whether the serum CA19-9 level is a significant predictor for survival and to determine the optimal cutoff value of CA19-9 for predicting prognosis.MethodsA total of 120 consecutive patients who underwent surgery for potentially resectable primary PDAC were retrospectively analyzed. The variables included the following: age, sex, the location of the tumor, the maximal tumor size, the histological differentiation, the margin status, the tumor stage, serum CA19-9 levels, and serum total bilirubin (TBil) levels.ResultsThe overall 1-year survival rate was 62.5%. The receiver operating characteristic (ROC) curve indicated a significant result for the level of CA19-9 in predicting death within 1 year after surgery (Area under the curve (AUC), 0.612; 95% confidence interval (CI), 0.505-0.720; P = 0.040). The optimal cutoff point was 338.45 U/mL (sensitivity, 60.0%; specificity, 66.7%; accuracy, 64.2%). The strongest univariate predictor among the categorized CA19-9 values was CA19-9 greater than or equal to 338.45 U/mL. In the multivariate Cox proportional hazards mode analysis, the serum CA19-9 level, age and the histological differentiation were significant independent prognostic factors that were associated with the overall survival.ConclusionsThe preoperative elevated CA19-9 level is a promising independent factor for predicting a poor prognosis in PDAC, and the optimal cutoff value is 338.45 U/mL.

GSTP1 determines cis-platinum cytotoxicity in gastric adenocarcinoma MGC803 cells: regulation by promoter methylation and extracellular regulated kinase signaling

Detoxification mechanisms can play a pivotal role in determining tumor cell responses to platinum-based chemotherapy. Glutathione S-transferase-pi (GSTP1) belongs to a supergene family of detoxifying enzymes involved in the prevention of DNA damage and subsequent platinum resistance in numerous cancers. The role of GSTP1 in gastric cancer sensitivity to chemotherapy is, however, not known. In this study, we found that the human gastric cancer cell line MGC803 was significantly more sensitive to cis-platinum (CDDP) than the other gastric cancer lines examined (BGC823 and SGC7901). To explore the potential role of GSTP1 in drug resistance, we measured GSTP1 expression in these cells. GSTP1 mRNA and protein were not detectable in MGC803 cells; both were present in BGC823 and SGC7901 cells. GSTP1 CpG island DNA methylation was examined. We report that promoter hypermethylation was associated with the absence of GSTP1 expression in MGC803 cells. Treatment of these cells with 5-aza-2′-deoxycytidine, a DNA methyltransferase inhibitor, restored GSTP1 expression and suppressed sensitivity to CDDP. The selective mitogen-activated protein kinase/extracellular regulated kinase (ERK) pathway inhibitor PD98059 decreased GSTP1 expression in 5-aza-2′-deoxycytidine-treated cells. A similar decrease was observed in the BGC823 and SGC7901 cell lines, suggesting that mitogen-activated protein kinase/ERK signaling stimulates GSTP1 expression. CDDP sensitivity was also enhanced by PD98059. These observations indicate that somatic promoter hypermethylation and impaired ERK signaling are associated with decreased GSTP1 expression and CDDP sensitivity in gastric cancer cell lines. Evaluation of promoter methylation and ERK activity may be useful for predicting tumor sensitivity to platinum-based chemotherapeutics.

c-Src expression is predictive of poor prognosis in breast cancer patients with bone metastasis, but not in patients with visceral metastasis

Zhang L, Teng Y, Zhang Y, Liu J, Xu L, Qu J, Hou K, Yang X, Liu Y, Qu X. c‐Src expression is predictive of poor prognosis in breast cancer patients with bone metastasis but not in patients with visceral metastasis. APMIS 2012; 120: 549–57.

MicroRNA-891b is an independent prognostic factor of pancreatic cancer by targeting Cbl-b to suppress the growth of pancreatic cancer cells

Growing evidence has revealed that microRNAs could regulate the proliferation of pancreatic ductal adenocarcinoma (PDAC) cells and predict the prognosis of PDAC. Here the comparative microRNA expression profiles of the good and poor prognosis groups were performed by microRNA microarray. MicroRNA-891b (miR-891b) was screened and validated to be a prognostic predictor of PDAC in the initial group and further evaluated to be an independent predictor for the overall survival of resectable PDACs in an independent cohort. By a series of cellular and animal experiments, as well as clinical specimen analyses, miR-891b was confirmed to target the Cbl-b gene, promot the expression of tumor suppressor p21 protein and inhibit the proliferation of PDAC cells. The results provide a theoretical basis for the study of miR-891b as an independent prognostic predictor of PDAC and the role of miR-891b/Cbl-b pathway in this prediction, as well as the identification of new targets for PDAC.