The Hippo pathway restricts the activity of transcriptional coactivators TAZ (WWTR1) and YAP. TAZ and YAP are reported to be overexpressed in various cancers, however, their prognostic significance in colorectal cancers remains unstudied. The expression levels of TAZ and YAP, and their downstream transcriptional targets, AXL and CTGF, were extracted from two independent colon cancer patient datasets available in the Gene Expression Omnibus database, totaling 522 patients. We found that mRNA expressions of both TAZ and YAP were positively correlated with those of AXL and CTGF (p<0.05). High level mRNA expression of TAZ, AXL or CTGF significantly correlated with shorter survival. Importantly, patients co-overexpressing all 3 genes had a significantly shorter survival time, and combinatorial expression of these 3 genes was an independent predictor for survival. The downstream target genes for TAZ-AXL-CTGF overexpression were identified by Java application MyStats. Interestingly, genes that are associated with colon cancer progression (ANTXR1, EFEMP2, SULF1, TAGLN, VCAN, ZEB1 and ZEB2) were upregulated in patients co-overexpressing TAZ-AXL-CTGF. This TAZ-AXL-CTGF gene expression signature (GES) was then applied to Connectivity Map to identify small molecules that could potentially be utilized to reverse this GES. Of the top 20 small molecules identified by connectivity map, amiloride (a potassium sparing diuretic,) and tretinoin (all-trans retinoic acid) have shown therapeutic promise in inhibition of colon cancer cell growth. Using MyStats, we found that low level expression of either ANO1 or SQLE were associated with a better prognosis in patients who co-overexpressed TAZ-AXL-CTGF, and that ANO1 was an independent predictor of survival together with TAZ-AXL-CTGF. Finally, we confirmed that TAZ regulates Axl, and plays an important role in clonogenicity and non-adherent growth in vitro and tumor formation in vivo. These data suggest that TAZ could be a therapeutic target for the treatment of colon cancer.

Purpose: Cancer cells have been shown to be more susceptible to Ran knockdown than normal cells. We now investigate whether Ran is a potential therapeutic target of cancers with frequently found mutations that lead to higher Ras/MEK/ERK [mitogen-activated protein/extracellular signal-regulated kinase (ERK; MEK)] and phosphoinositide 3-kinase (PI3K)/Akt/mTORC1 activities.Experimental Design: Apoptosis was measured by flow cytometry [propidium iodide (PI) and Annexin V staining] and MTT assay in cancer cells grown under different conditions after knockdown of Ran. The correlations between Ran expression and patient survival were examined in breast and lung cancers.Results: Cancer cells with their PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways inhibited are less susceptible to Ran silencing-induced apoptosis. K-Ras-mutated, c-Met-amplified, and Pten-deleted cancer cells are also more susceptible to Ran silencing-induced apoptosis than their wild-type counterparts and this effect is reduced by inhibitors of the PI3K/Akt/mTORC1 and MEK/ERK pathways. Overexpression of Ran in clinical specimens is significantly associated with poor patient outcome in both breast and lung cancers. This association is dramatically enhanced in cancers with increased c-Met or osteopontin expression, or with oncogenic mutations of K-Ras or PIK3CA, all of which are mutations that potentially correlate with activation of the PI3K/Akt/mTORC1 and/or Ras/MEK/ERK pathways. Silencing Ran also results in dysregulation of nucleocytoplasmic transport of transcription factors and downregulation of Mcl-1 expression, at the transcriptional level, which are reversed by inhibitors of the PI3K/Akt/mTORC1 and MEK/ERK pathways.Conclusion: Ran is a potential therapeutic target for treatment of cancers with mutations/changes of expression in protooncogenes that lead to activation of the PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways. Clin Cancer Res; 18(2); 380-91. Ó2011 AACR.

Our results suggest that upregulation of Twist plays a role in the neoplastic transformation to oesophageal SCC and subsequent development of distant metastasis. Twist may serve as a useful prognostic marker for predicting the development of distant metastasis in oesophageal SCC.

Recent studies have revealed an oncogenic role of DJ-1 through its ability to transform normal cells, prevent oxidative damage, and inhibit apoptosis. However, its role in esophageal squamous cell carcinoma (ESCC) is unknown. In this study, by immunohistochemistry, we analyzed the expression of DJ-1 in 81 ESCC tumors, 31 paired nonneoplastic esophageal epithelia, and 19 paired ESCC lymph node metastases. We found that cytoplasmic DJ-1 expression was significantly higher in ESCC and ESCC lymph node metastases than in nonneoplastic esophageal epithelium. ESCC specimens with high distant metastatic potential also had a significantly higher level of nuclear DJ-1 expression (P = 0.018). By Kaplan-Meier analysis, we found that a high level of nuclear DJ-1 was significantly associated with poorer patient survival in our cohort (P = 0.028). To investigate whether DJ-1 promotes ESCC progression through phosphatidylinositol 3-kinase pathway and modulation of apoptosis, we performed immunohistochemistry of pAkt and Daxx. We found that DJ-1 expression was significantly associated with pAkt, whereas nuclear DJ-1 expression was significantly correlated with nuclear expression of Daxx. These results suggest that phosphatidylinositol 3-kinase pathway and Daxx-regulated apoptosis might be important in DJ-1-mediated ESCC progression. By using multivariate Cox regression, we further showed that T 4 stage (P = 0.003) and DJ-1 (P = 0.034) are independent predictors of patient survival. In conclusion, our results suggest that DJ-1 plays a very important role in transformation and progression of ESCC and may be used as a prognostic marker in ESCC. (Cancer Epidemiol Biomarkers Prev 2008;17(12):3593 -602)

Our results suggest that Ran is required for and is a potential therapeutic target of Myc-driven cancer progression in both breast and lung cancers.

The core LATS kinases of the Hippo tumor suppressor pathway phosphorylate and inhibit the downstream transcriptional coactivators YAP and TAZ, which are implicated in various cancers. Recent studies have identified various E3 ubiquitin ligases that negatively regulate the Hippo pathway via ubiquitination, yet few deubiquitinating enzymes (DUB) have been implicated. In this study, we report the DUB USP9X is an important regulator of the core kinases of this pathway. USP9X interacted strongly with LATS kinase and to a lesser extent with WW45, KIBRA, and Angiomotin, and LATS co-migrated exclusively with USP9X during gel filtration chromatography analysis. Knockdown of USP9X significantly downregulated and destabilized LATS and resulted in enhanced nuclear translocation of YAP and TAZ, accompanied with activation of their target genes. In the absence of USP9X, cells exhibited an epithelial-to-mesenchymal transition phenotype, acquired anchorage-independent growth in soft agar, and led to enlarged, disorganized, three-dimensional acini. YAP/TAZ target gene activation in response to USP9X knockdown was suppressed by knockdown of YAP, TAZ, and TEAD2. Deletion of USP9X in mouse embryonic fibroblasts resulted in significant downregulation of LATS. Furthermore, USP9X protein expression correlated positively with LATS but negatively with YAP/TAZ in pancreatic cancer tissues as well as pancreatic and breast cancer cell lines. Overall, these results strongly indicate that USP9X potentiates LATS kinase to suppress tumor growth. Cancer Res; 77(18); 4921-33. Ó2017 AACR.

Purpose: Previously, FTY720 was found to possess potent anticancer effects on various types of cancer. In the present study, we aimed to first verify the role of Runx2 in prostate cancer progression and metastasis, and, subsequently, assessed if FTY720 could modulate Runx2 expression, thus interfering downstream events regulated by this protein.Experimental Design: First, the association between Runx2 and prostate cancer progression was assessed using localized prostate cancer specimens and mechanistic investigation of Runx2-induced cancer aggressiveness was then carried out. Subsequently, the effect of FTY720 on Runx2 expression and transcriptional activity was investigated using PC-3 cells, which highly expressed Runx2 protein. Last, the involvement of Runx2 in FTY720-induced anticancer effects was evaluated by modulating Runx2 expression in various prostate cancer cell lines. Results: Runx2 nuclear expression was found to be up-regulated in prostate cancer and its expression could be used as a predictor of metastasis in prostate cancer. Further mechanistic studies indicated that Runx2 accelerated prostate cancer aggressiveness through promotion of cadherin switching, invasion toward collagen I, and Akt activation. Subsequently, we found that FTY720 treatment down-regulated Runx2 expression and its transcriptional activity, as well as inhibited its regulated downstream events. More importantly, silencing Runx2 in PC-3 enhanced FTY720-induced anticancer effects as well as cell viability inhibition, whereas overexpressing Runx2 in 22Rv1that expressed very low endogenous Runx2 protein conferred resistance in the same events. Conclusion: This study provided a novel mechanism for the anticancer effect of FTY720 on advanced prostate cancer, thus highlighting the therapeutic potential of this drug in treating this disease.

A major cause of treatment failure for prostate cancer is the development of androgen-independent metastatic disease. Id protein family, a group of basic helix-loop-helix transcription factors, has been shown to be involved in carcinogenesis and a prognostic marker in several types of human cancers. In this study, we examined the expressions of four Id proteins, Id-1, -2, -3 and -4, in 125 clinical prostate cancer specimens as well as 40 nodular hyperplasia specimens by immunohistochemistry. The expressions of Id proteins were correlated with Gleason grading and metastatic progress of the tumors. We found that Id proteins were dysregulated in prostate cancer. Id-1 and -2 expressions were elevated while Id-3 and -4 expressions were reduced in prostate cancers compared to nodular hyperplasia. Cytoplasmic staining of Id-1 (P ¼ 0.013) and nuclear staining of Id-2 (P ¼ 0.001) and Id-4 (Po0.001) were positively correlated with Gleason score. The results indicate that these Id proteins may play a positive role in the development of prostate cancer. In contrast, Id-3 might have an inverse relationship with prostate neoplastic transformation (P ¼ 0.002) and cancer progression (P ¼ 0.022). We found that Id-4 nuclear overexpression in the primary prostate cancers significantly increased the risks to the development of metastasis in the patients (odds ratio ¼ 3.215, 95% confidence interval ¼ 1.150-8.987, P ¼ 0.026). Our results suggest that in prostate cancer patients, differential Id proteins expressions may be a useful marker for poor prognosis, and Id-4 may be a potential prognostic marker for distant metastasis.