Dishui Gu

Gas6/Axl Axis Contributes to Chemoresistance and Metastasis in Breast Cancer through Akt/GSK-3β/β-catenin Signaling

Chemoresistance in breast cancer has been of great interest in past studies. However, the development of rational therapeutic strategies targeting chemoresistant cells is still a challenge in clinical oncology. By integrating data from global differences of gene expression and phospho-receptor tyrosine kinases between sensitive parental cells (MCF-7) and doxorubicin-resistant cells (MCF-7/ADR), we identified Axl as a potential target for chemoresistance and metastasis in multidrug resistant breast cancer cells. We analyzed Axl expression in 57 breast cancer cell lines and detected a dramatic increase in its expression level in mesenchymal breast cancer cell lines. Axl silencing suppressed invasive and metastatic potentials of chemoresistant breast cancer cells as well as increased elimination of cancer cells when combined with doxorubicin. Furthermore, in preclinical assays, an Axl inhibitor R428 showed increased cell death upon doxorubicin treatment. Additionally, using phospho-kinase array based proteomic analysis, we identified that Akt/GSK-3β/β-catenin cascade was responsible for Axl-induced cell invasion. Nuclear translocation of β-catenin then induced transcriptional upregulation of ZEB1, which in turn regulated DNA damage repair and doxorubicin-resistance in breast cancer cells. Most importantly, Axl was correlated with its downstream targets in tumor samples and was associated with poor prognosis in breast cancer patients. These results demonstrate that Gas6/Axl axis confers aggressiveness in breast cancer and may represent a therapeutic target for chemoresistance and metastasis.

LIFR functions as a metastasis suppressor in hepatocellular carcinoma by negatively regulating phosphoinositide 3-kinase/AKT pathway

Hepatocellular carcinoma (HCC) is one of the leading causes for cancer related mortality worldwide. Poor prognosis of HCC patients is mainly due to frequent metastasis and recurrence. Deregulation of metastasis suppressors in malignant cells plays critical roles during cancer metastasis. Thus, novel metastasis suppressors are urgently needed to be uncovered to shed new light on molecular mechanisms driving HCC metastasis. In the present study, decreased expression of leukemia inhibitory factor receptor (LIFR) was demonstrated in HCC, and its expression levels were even lower in HCC with metastasis. Downregulated LIFR expression predicted poor prognosis in HCC patients. LIFR was an independent and significant risk factor for their recurrence and survival. Silencing LIFR resulted in forced metastasis of HCC cells, whereas ectopic overexpression of LIFR attenuated migration and invasion of HCC cells in vitro and in vivo. Moreover, LIFR knockdown could activate phosphoinositide 3-kinase/V-akt Murine Thymoma Viral Oncogene Homolog (PI3K/AKT) signaling through enhancing phosphorylation of Janus kinase 1 (JAK1), which successively promoted matrix metalloproteinase 13 (MMP13) expression and HCC metastasis. Combination of LIFR and p-AKT or MMP13 was a more powerful predictor of poor prognosis for HCC patients. Together, these findings conclude that LIFR functions as a novel metastasis suppressor in HCC and may serve as a prognostic biomarker for HCC patients.

Prognostic significance of kynurenine 3-monooxygenase and effects on proliferation, migration, and invasion of human hepatocellular carcinoma

Kynurenine 3-monooxygenase (KMO) is a pivotal enzyme in the kynurenine pathway of tryptophan degradation and plays a critical role in Huntington’s and Alzheimer’s diseases. This study aimed to examine the expression of KMO in human hepatocellular carcinoma (HCC) and investigate the relationship between its expression and prognosis of HCC patients. We first analyzed KMO expression in 120 paired HCC samples (HCC tissues vs matched adjacent non-cancerous liver tissues), and 205 clinical HCC specimens using immunohistochemistry (IHC). Kaplan-Meier survival and Cox regression analyses were executed to evaluate the prognosis of HCC. The results of IHC analysis showed that KMO expression was significantly higher in HCC tissues than that in normal liver tissues (all p < 0.05). Survival and recurrence analyses showed that KMO was an independent prognostic factor for overall survival (OS) and time to recurrence (TTR) (both p<0.01). And in vitro studies revealed that KMO positively regulated proliferation, migration, and invasion of HCC cells. These results suggest that KMO exhibits tumor-promoting effects towards HCC and it may serve as a novel prognostic marker in HCC.

KDM6B Elicits Cell Apoptosis by Promoting Nuclear Translocation of FOXO1 in Non-Small Cell Lung Cancer

Background/Aims: Non-small cell lung carcinoma (NSCLC) is the most common type of lung cancer and the cause of most cancer-related deaths. The molecular mechanisms that are involved in NSCLC development are currently not well understood. Accumulating evidence shows that histone demethylases play important roles in the regulation of pathological developmental processes in many diseases, including various types of cancers. Methods: Mitochondrial membrane potential assays, migration and invasion assays, caspase-3 and caspase-9 activity assays and western blot analysis were used in this research. Results: We found that overexpression of KDM6B, a demethylase that acts on histone H3 at lysine 27 (H3K27), inhibited cell growth by initiating mitochondria-dependent apoptosis and by attenuating the invasion-metastasis cascade in NSCLC cells. Moreover, our results showed that KDM6B directly interacted with FOXO1 and that overexpression of KDM6B promoted nuclear accumulation of FOXO1. The effects of KDM6B on cell apoptosis and metastasis were weakened by knockdown of FOXO1 expression. On the contrary, knocking down expression of KDM6B inhibited cell apoptosis and promoted cell growth by mitigating the nuclear translocation of FOXO1 in NSCLC cells. Conclusions: These findings suggest that KDM6B may act in a pro-apoptotic role in NSCLC by causing the nuclear translocation of FOXO1.

Long noncoding RNA SchLAH suppresses metastasis of hepatocellular carcinoma through interacting with fused in sarcoma

Emerging evidence has indicated that deregulation of long non‐coding RNAs (lncRNAs) can contribute to the progression and metastasis of human cancer, including hepatocellular carcinoma (HCC). However, the roles of most lncRNAs in HCC remain largely unknown. Here we found a long noncoding RNA termed SchLAH (seven chromosome locus associated with HCC; also called BC035072) was generally downregulated in HCC. Low expression of SchLAH was significantly correlated with shorter overall survival of HCC patients. In vitro and in vivo assays indicated that overexpression of SchLAH inhibited the migration and lung metastasis of HCC cells. Knockdown of SchLAH by siRNA pool promoted the migration of HCC cells. RNA pull‐down and RNA immunoprecipitation assays demonstrated SchLAH physically interacted with fused in sarcoma (FUS). PCR array analysis showed that RhoA and Rac1 were the downstream effector molecules of SchLAH during HCC metastasis. Knockdown of FUS rescued the mRNA levels of RhoA and Rac1 that were repressed by SchLAH. These results suggest that SchLAH may suppress the metastasis of HCC cells by interacting with FUS, which indicates potential of SchLAH for the prognosis and treatment of HCC.

The asialoglycoprotein receptor suppresses the metastasis of hepatocellular carcinoma via LASS2-mediated inhibition of V-ATPase activity

The asialoglycoprotein receptor (ASGR), which is expressed mainly in hepatocytes, is downregulated in poorly differentiated hepatocellular carcinoma (HCC). Here we investigated the role of ASGR1 in HCC metastasis as well as the possible underlying molecular mechanisms. We found that ASGR1 was downregulated in HCC tissue compared with adjacent non-tumorous liver tissue and that lower ASGR1 expression was associated with higher TNM stage and poorer prognosis in HCC patients. ASGR1 overexpression inhibited hepatoma cell migration and invasion in vitro and in vivo, while ASGR1 knockdown had the opposite effects. Furthermore, ASGR1 interacted directly with human longevity assurance homolog 2 of yeast LAG1 (LASS2). Knockdown of LASS2 attenuated the inhibitory effects of ASGR1 on hepatoma cell migration and invasion in vitro. ASGR1 decreased V-ATPase activity in hepatoma cells, and this was reversed by LASS2 knockdown. Finally, HCC patients with low LASS2 levels had poor prognosis, while those with high ASGR1 and LASS2 levels had better prognosis. Thus, ASGR1 may act as a potential metastasis suppressor in HCC, and the combination of ASGR1 and LASS2 may help predict the prognosis of HCC patients.

Liver-specific deletion of LASS2 delayed regeneration of mouse liver after partial hepatectomy.

The capacity of liver regeneration is critical for patients with liver diseases. However, cellular and molecular mechanisms of liver regeneration are still incompletely defined. Here, we assessed roles of LASS2 in liver regeneration following partial hepatectomy (PHx) in mice. Our results showed that protein level of LASS2 remarkably increased during liver regeneration after PHx in wildtype (WT) mice. Comparing to WT mice, liver regeneration index after PHx was significantly decreased from day 1 to day 5 in liver-specific LASS2 knockout (LASS2-LKO) mice. Interestingly, liver mass of LASS2-LKO mice could sufficiently recover at day 14 after PHx. Immunohistochemistry (IHC) and western blot analyses revealed that proliferation markers, such as PCNA and Ki67, were potently reduced during liver regeneration in LASS2-LKO mice. In addition, several cell cycle related molecules, such as cyclin A, CDK2 and p-Rb, were decreased in LASS2-LKO mice after PHx. Co-immunoprecipitation assay further revealed a decreased formation of CDK4/cyclin D1 complex after PHx in LASS2-LKO mice. However, phosphorylation of Akt was significantly activated from day 2 after PHx in LASS2-LKO mice when compared with that in WT mice, which may explain the recovery of liver mass at the late stage of liver regeneration in LASS2-LKO mice. Taken together, we conclude that LASS2 plays an important role in efficient liver regeneration in response to PHx.

Co-expression of LASS2 and TGF-β1 predicts poor prognosis in hepatocellular carcinoma

Longevity assurance homolog 2 of yeast LAG1 (LASS2) has been reported to act as an important tumor suppressor in the development of human cancers. However, little is known about the prognostic value of LASS2 in hepatocellular carcinoma (HCC) . In the present study, we analyzed correlation between LASS2 and TGF-β1 levels, and evaluated their prognostic values in HCC patients. We first analyzed the expression of LASS2 and TGF-β1 in two independent cohorts (test cohort: 184 HCC patients; validation cohort: 118 HCC patients) using immunohistochemistry (IHC). Kaplan-Meier survival and Cox regression analyses were executed to evaluate the prognosis of HCC. The results of IHC analysis revealed a positive correlation between the expression of LASS2 and TGF-β1. HCC Patients with low expression of LASS2 and TGF-β1 had shorter overall survival (OS) and time to recurrence (TTR) than patients with high expression of LASS2 and TGF-β1. Furthermore, combination of LASS2 and TGF-β1 was an independent and significant risk factor for OS and TTR. In conclusion, low expression of LASS2 and TGF-β1 contributes to the aggressiveness and poor prognosis of HCC, and may represent a novel prognostic biomarker for HCC patients.