Puxiongzhi Wang

The molecular effect of metastasis suppressors on Src signaling and tumorigenesis: new therapeutic targets

A major problem for cancer patients is the metastasis of cancer cells from the primary tumor. This involves: (1) migration through the basement membrane; (2) dissemination via the circulatory system; and (3) invasion into a secondary site. Metastasis suppressors, by definition, inhibit metastasis at any step of the metastatic cascade. Notably, Src is a non-receptor, cytoplasmic, tyrosine kinase, which becomes aberrantly activated in many cancer-types following stimulation of plasma membrane receptors (e.g., receptor tyrosine kinases and integrins). There is evidence of a prominent role of Src in tumor progression-related events such as the epithelial–mesenchymal transition (EMT) and the development of metastasis. However, the precise molecular interactions of Src with metastasis suppressors remain unclear. Herein, we review known metastasis suppressors and summarize recent advances in understanding the mechanisms of how these proteins inhibit metastasis through modulation of Src. Particular emphasis is bestowed on the potent metastasis suppressor, N-myc downstream regulated gene 1 (NDRG1) and its interactions with the Src signaling cascade. Recent studies demonstrated a novel mechanism through which NDRG1 plays a significant role in regulating cancer cell migration by inhibiting Src activity. Moreover, we discuss the rationale for targeting metastasis suppressor genes as a sound therapeutic modality, and we review several examples from the literature where such strategies show promise. Collectively, this review summarizes the essential interactions of metastasis suppressors with Src and their effects on progression of cancer metastasis. Moreover, interesting unresolved issues regarding these proteins as well as their potential as therapeutic targets are also discussed.

Tumor-derived CXCL5 promotes human colorectal cancer metastasis through activation of the ERK/Elk-1/Snail and AKT/GSK3β/β-catenin pathways

BackgroundMetastasis is a major cause of death in human colorectal cancer patients. However, the contribution of chemokines in the tumor microenvironment to tumor metastasis is not fully understood.MethodsHerein, we examinined several chemokines in colorectal cancer patients using chemokine ELISA array. Immunohistochemistry was used to detect expression of CXCL5 in colorectal cancer patients tissues. Human HCT116 and SW480 cell lines stably transfected with CXCL5, shCXCL5 and shCXCR2 lentivirus plasmids were used in our in vitro study. Immunoblot, immunofluorescence and transwell assay were used to examine the molecular biology and morphological changes in these cells. In addition, we used nude mice to detect the influence of CXCL5 on tumor metastasis in vivo.ResultsWe found that CXCL5 was overexpressed in tumor tissues and associated with advanced tumor stage as well as poor prognosis in colorectal cancer patients. We also demonstrated that CXCL5 was primarily expressed in the tumor cell cytoplasm and cell membranes, which may indicate that the CXCL5 was predominantly produced by cancer epithelial cells instead of fibroblasts in the tumor mesenchyme. Additionally, overexpression of CXCL5 enhanced the migration and invasion of colorectal cancer cells by inducing the epithelial-mesenchymal transition (EMT) through activation of the ERK/Elk-1/Snail pathway and the AKT/GSK3β/β-catenin pathway in a CXCR2-dependent manner. The silencing of Snail and β-catenin attenuated CXCL5/CXCR2-enhanced cell migration and invasion in vitro. The elevated expression of CXCL5 can also potentiate the metastasis of colorectal cancer cells to the liver in vivo in nude mice intrasplenic injection model.ConclusionIn conclusion, our findings support CXCL5 as a promoter of colorectal cancer metastasis and a predictor of poor clinical outcomes in colorectal cancer patients.

Cadherin-12 contributes to tumorigenicity in colorectal cancer by promoting migration, invasion, adhersion and angiogenesis

BackgroundCadherin 12 (CDH12), which encodes a type II classical cadherin from the cadherin superfamily, may mediate calcium-dependent cell adhesion. It has been demonstrated that CDH12 could play an important role in the invasion and metastasis of salivary adenoid cystic carcinoma. We decided to investigate the relationship between CDH12 expression level and clinicopathologic variables in colorectal carcinoma (CRC) patients and to explore the functions of CDH12 in tumorigenesis in CRC.MethodsThe expression levels of CDH12 in colorectal carcinoma tissues were detected by immunohistochemistry. Real-time PCR and Western Blot were used to screen CDH12 high-expression cell lines. CCK-8 assay was used to detect the proliferation ability of CRC cells being transfected by shRNAs against CDH12. The wound assay and transwell assay were performed to test migration and invasion ability. The importance of CDH12 in cell-cell junctions was detected by cell adhesion assay and cell aggregation assay. Endothelial tube formation assay was used to test the influence of CDH12 on angiogenesis.ResultsStatistical analysis of clinical cases revealed that the positive rate of CDH12 was higher in the CRC tumor tissues compared with the adjacent non-tumor tissues. The expression levels of CDH12 in CRC patients are significantly correlated with invasion depth. Consistently, the ability of proliferation, migration and invasion were suppressed when CDH12 was decreased in CRC cells transfected with shRNAs. Cell adhesion assay and cell aggregation assay presented that tumor cells tend to disperse with the lack of CDH12. Endothelial tube formation assay showed that down-regulation of CDH12 could obviously inhibit the process of angiogenesis, implying that CDH12 may play an important role in tumor metastasisConclusionOur results showed that CDH12 promotes proliferation, migration, invasion, adhesion and angiogenesis, suggesting that CDH12 may be an oncogene in colorectal cancer. CDH12 is expected to become a new diagnostic and prognostic marker and a novel target of the treatment of colorectal cancer.

Cadherin-12 enhances proliferation in colorectal cancer cells and increases progression by promoting EMT

Cadherin-12 (CDH12) is a subtype of N-cadherin family. In this study, we investigated the expression of CDH12 and the role of CDH12 in prognosis of colorectal cancer (CRC) patients. In addition, we observed the influence of CDH12 on proliferation and progression of CRC cell lines. By using immunohistochemical staining, we analyzed CRC samples and adjacent non-tumor tissues collected from 78 patients who underwent laparoscopic surgery in Shanghai Minimally Invasive Center, China. Statistical analyses were used to analyze relationship between CDH12 and tumor features. Kaplan-Meier method was used to analyze patients’ survival. Proliferation ability of CRC cells was tested by CCK-8 assay, and transwell assays were performed to detect migration and invasion ability. Western blot assay was performed to investigate epithelial-mesenchymal transition (EMT) variants. We found that expression of CDH12 in tumor tissue was higher than in adjacent normal tissue. High expression of CDH12 was associated with tumor invasion depth and predicts poor prognosis of CRC patients. Ectopic/repressing expression of CDH12 increased/decreased the proliferation and migration ability of CRC cells. CDH12 is able to increase cancer cell migration and invasion via promoting EMT by targeting transcriptional factor Snail. These findings may conclude that CDH12 may act as a predictor in CRC patients’ prognosis and an oncogene in CRC cell proliferation and migration. CDH12 may influence CRC cell progression through promoting EMT by targeting Snail. In addition, CDH12 is promoted by MCP1 through induction of MCPIP.

PFDN1, an indicator for colorectal cancer prognosis, enhances tumor cell proliferation and motility through cytoskeletal reorganization.

Prefoldin (PFDN) subunits have been reported upregulated in various tumor types, while the expression and functions of PFDN1 (PFDN subunit 1) in colorectal cancer (CRC) are not well elucidated. The aim of this study was to investigate the use of PFDN1 as a poor prognosis indicator for CRC and explore the functions of PFDN1 in CRC. The relationship between PFDN1 expression and CRC clinical-pathological statistics was detected on the tissue microarray containing 145 cases of CRC. ShRNA was used to silence PFDN1 expression in SW480 and RKO CRC cells, and these transfected cells were analyzed for changes in proliferation, colony formation, cell cycle, migration, and invasion. Immunofluorescence and immunoblot were used to determine the remodeling of the F-actin and α-tubulin. Finally, tumor growth on nude mice was observed and measured. In this study, we found PFDN1 was upregulated in CRC tissues compared with adjacent normal tissues. Also, PFDN1 expression positively correlated with tumor size and tumor invasion. Moreover, after silencing PFDN1 in SW480 and RKO cells, the proliferation and motility of CRC cells were significantly suppressed. The inhibitory effect of PFDN1 on tumor cell growth and motility was partially due to G2/M cell cycle blockage and cytoskeletal deficiency. Finally, in vivo assay showed that downregulation of PFDN1 inhibited tumor growth on nude mice and PFDN1 expression correlated with higher levels of Ki-67 staining. These findings indicate that PFDN1 was involved in the progression of CRC, and provide new insights into PFDN1 as a potential therapeutic target for CRC treatment.

Overexpression of CXCR2 predicts poor prognosis in patients with colorectal cancer

Colorectal cancer is a heterogeneous disease. Although many risk factors are used to predict colorectal cancer patients’ prognosis after surgical resection, new prognostic factors are still needed to be defined to promote predictive efficacy of prognosis and further guide therapies. Herein, we identified the prognostic significance of CXCR2 in colorectal cancer patients. We retrospectively analysed 134 patients with colorectal cancer who underwent minimally invasive surgery between 2010 and 2011. The overall cohort was divided into a training set (n = 78) and a validation set (n = 56). We detected CXCR2 expression using immunohistochemical staining and defined the cut-off value using X-tile program. Next, we analysed the association between CXCR2 expression and clinicopathologic features in training and validation sets. High expression of CXCR2 was associated with Dukes stage (P = 0.018), tumor invasion (P = 0.018) and liver metastasis (P = 0.047). Multivariate COX regression analyses confirmed that high CXCR2 level was an independent prognostic risk factor for both overall survival and disease free survival. Kaplan-Meier survival analysis demonstrated that patients with high expression of CXCR2 had a poor overall survival and disease free survival even in low-risk group (I + II). This indicated that CXCR2 can help to refine individual risk stratification. In addition, we established Nomograms of all significant factors to predict 3- or 5-years overall survival and disease free survival. Moreover, we found the combination of CXCR2 and its ligand CXCL5 had more significant value in predicting the prognosis than single CXCR2 factor.

Bone marrow-derived mesenchymal stromal cells promote colorectal cancer cell death under low-dose irradiation

Background:Radiotherapy remains one of the cornerstones to improve the outcome of colorectal cancer (CRC) patients. Radiotherapy of the CRC not only help to destroy cancer cells but also remodel the tumour microenvironment by enhancing tumour-specific tropism of bone marrow-derived mesenchymal stromal cell (BM-MSC) from the peripheral circulation. However, the role of local MSCs and recruited BM-MSC under radiation were not well defined. Indeed, the functions of BM-MSC without irradiation intervention remained controversial in tumour progression: BM-MSC was previously shown to modulate the immune function of major immune cells, resulting in an impaired immunological sensitivity and to induce an increased risk of tumour recurrence. In contrast, it could also secrete various cytokines and possess anticancer effect.Methods:Three co-cultivation modules, 3D culture modules, and cancer organoids were established. The induction of cytokines secretion in hBM-MSCs after irradiation was analysed by ELISA array and flow cytometry. AutoMac separator was used to separate hBM-MSC and CRC automatically. Cells from the co-cultured group and the control group were then irradiated by UV-C lamp and X-ray. Proliferation assay and viability assay were performed.Results:In this study, we show that BM-MSCs can induce the EMT progression of CRC cells in vitro. When irradiated with low doses of ultraviolet radiation and X-rays, BM-MSCs show an anti-tumour effect by secreting certain cytokine (TNF-α, IFN-γ) that lead to the inhibition of proliferation and induction of apoptosis of CRC cells. This was further verified in a 3D culture model of a CRC cell in vitro. Furthermore, irradiation on the co-culture system induced the cleavage of caspase3, and attenuated the phosphorylation of phosphatidylinositol 3-kinase (PI3K)/AKT and extracellular signal-regulated kinase in cancer cells. The signal pathways above might contribute to the cancer cell death.Conclusions:Taken together, we show that BM-MSC can potentially promote the effect of radiotherapy in CRC.