Mortalin maintains breast cancer stem cells stemness via activation of Wnt/GSK3β/β-catenin signaling pathway

Previous research indicated that mortalin overexpressed in breast cancer and contributed to carcinogenesis. Mortalin was also demonstrated to promote Epithelial-mesenchymal transition (EMT) and was considered as a factor for maintaining the stemness of the cancer stem cells. However, the underlying mechanisms about mortalin maintaining the stemness of breast cancer stem cells (BCSCs) remain unclear. Here, we identified that increased expression of mortalin in breast cancer was associated with poorer overall survival rate. Mortalin was elevated in breast cancer cell lines and BCSC-enriched populations. Additionally, knockdown of mortalin significantly inhibited the cell proliferation, migration and EMT, as well as sphere forming capacity and stemness genes expression. Further study revealed that mortalin promoted EMT and maintained BCSCs stemness via activating the Wnt/GSK3β/β-catenin signaling pathway in vivo and in vitro. Taken together, these findings unveiled the mechanism of mortalin in maintaining and regulating the stemness of BCSCs, and may offer novel therapeutic strategies for breast cancer treatment.     

Targeting cancer stem cells via integrin β4

Integrins mediate cell-cell interactions and communication with the extracellular matrix (ECM). These transmembrane protein receptors allow binding between a cell and its surroundings, initiating a breadth of intracellular signaling resulting in proliferation, differentiation, survival, or migration. Such responses have made integrins an attractive target for cancer therapy. Self-renewing and highly tumorigenic cancer stem cells (CSCs) are most resistant to traditional radiation treatment and chemotherapy, and therefore may contribute directly to the metastasis and relapse of the disease. In both the 4T1 mouse metastatic mammary tumor model and SCC7 head and neck squamous cell carcinoma model, integrin β4 (ITGB4) was expressed on ALDH^high 4T1 and SCC7 CSCs. Using two immunological approaches, we targeted ITGB4 through 1) ITGB4 protein-pulsed dendritic cell (ITGB4-DC) vaccination or 2) via anti-CD3/anit-ITGB4 bispecific antibody (ITGB4 BiAb)-armed T cell adoptive transfer. These two therapies reduced ITGB4-expressing CSCs and inhibited local tumor growth and lung metastasis through ITGB4 specific cellular and humoral immune responses. Additionally, the combination of anti-PD-L1 immunotherapy with our two ITGB4-targeted approaches significantly improved treatment efficacy. We also found increased concentrations of serum IFN-γ and IL-6 in the 4T1 and SCC7 models which may help define future directions of this ITGB4-targeted study. Together, these results emphasize ITGB4 as a practical CSC immunological target with possible therapeutic benefits across tumor types with high ITGB4 expression.     

Inhibition of Dishevelled-2 suppresses the biological behavior of pancreatic cancer by downregulating Wnt/β-catenin signaling

Dishevelled-2 (DVL2) has been proven to be involved in the tumorigenesis of several human cancers, such as colorectal cancer, lung cancer, prostate cancer, etc. However, its role in pancreatic ductal adenocarcinoma (PDAC) remains unclear. The present study investigated the effects of aberrantly expressed DVL2 on PDAC. A total of 97 pancreatic cancer (PC) samples and 85 adjacent normal samples were obtained from patients who were histopathologically diagnosed with primary PDAC. The present study demonstrated that DVL2 expression was upregulated in PDAC tissues and was positively associated with advanced clinical stage and lymph node metastasis in patients with PDAC. In addition, patients with high expression of DVL2 had a shorter overall survival rate compared with those with low expression. To elucidate the role of DVL2 in PDAC, lentivirus-mediated short hairpin RNA was used to silence DVL2 and its physiological function was analyzed in CFPAC-1 and PANC-1 cells. The results indicated that DVL2 downregulation significantly impaired its oncogenic functions including cell proliferation, migration, invasion and epithelial-mesenchymal transition. Furthermore, DVL2 knockdown inhibits the proliferation and invasion of PC cells in vivo. In addition, co-immunoprecipitation assays revealed that DVL2 interacted with β-catenin; knockdown of DVL2 reduced the expression level of β-catenin and inhibited β-catenin translocation into the nucleus. In conclusion the findings of the present study suggested that DVL2 may be a potential therapeutic target in the treatment of PDAC.     

Knockdown of Obg-like ATPase 1 enhances sorafenib sensitivity by inhibition of GSK-3β/β-catenin signaling in hepatocellular carcinoma cells

BackgroundTo clarify the molecular mechanism of hepatocellular carcinoma (HCC), conducive to developing an effective HCC therapy. Owing to the severe drug resistance, the clinical use of sorafenib, which is approved for HCC treatment, is limited. The precise molecular mechanisms of sorafenib drug resistance remain unclear. In the current work, we evaluated the role of Obg-like ATPase 1 (OLA1) in sorafenib resistance in HCC.MethodsThe survival of HCC patients between OLA1 expression and sorafenib treatment was analyzed by Kaplan-Meier plotter. Cell viability was measured by cell counting kit-8 (CCK-8) and colony formation assays. Cell death was detected by propidium iodide (PI) and trypan blue staining. The mRNA and protein levels were measured by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot (WB), respectively.ResultsWe found that OLA1 was highly correlated with sorafenib resistance of HCC through a public database. Further study showed that knockdown of OLA1 enhanced cell proliferation inhibition and cell death induced by sorafenib, along with a reduction of proliferation-associated proteins (c-Myc and cyclin D1) and increase of apoptosis-related proteins (cleaved caspase-3 and cleaved PARP) in HCC cells. In addition, knockdown of OLA1 reduced the activation of glycogen synthase kinase 3β (GSK-3β)/β-catenin.ConclusionsOur results proved that OLA1 can be a potential target to enhance sorafenib sensitivity in HCC.     

Dihydroartemisinin suppresses proliferation,migration, the Wnt/β-catenin pathway and EMT via TNKS in gastric cancer

Gastric cancer is a common malignancy worldwide. However, the molecular mechanisms underlying this malignancy remain unclear and there are a lack of effective drugs. The present study aimed to investigate the antitumor effect of Dihydroartemisinin (DHA) or inhibition of Tankyrases (TNKS), and determine the underlying molecular mechanisms of gastric cancer. Immunohistochemistry and immunofluorescence analyses were performed to detect the expression levels of TNKS, epithelial-to-mesenchymal transition (EMT) and Wnt/β-catenin pathway-related proteins in gastric cancer tissues and adjacent normal tissues. The Cell Counting Kit-8 assay was performed to assess the viability of HGC-27 and AGS cells following treatment with different concentrations of HLY78 (a Wnt activator) or DHA. Following treatment with HLY78, DHA or small interfering (si)-TNKS1/si-TNKS2, colony formation and migratory abilities were assessed via the colony formation, wound healing and Transwell assays. Furthermore, western blot and immunofluorescence analyses were performed to detect the expression levels of TNKS, EMT- and Wnt/β-catenin-related proteins. The results demonstrated that the expression levels of TNKS, AXI2, β-catenin, N-cadherin and Vimentin were upregulated, whereas E-cadherin expression was downregulated in gastric cancer tissues compared with normal tissues. Furthermore, HLY78 and DHA suppressed the viability of HGC-27 and AGS cells, in a concentration-independent manner. Notably, TNKS knockdown or treatment with DHA suppressed colony formation, migration, TNKS expression, EMT and the Wnt/β-catenin pathway. Opposing effects were observed following treatment with HLY78, which were ameliorated following co-treatment with DHA. Taken together, these results suggest that DHA or inhibition of TNKS can suppress the proliferation and migration of gastric cancer cells, which is partly associated with inactivation of the Wnt/β-catenin pathway and EMT process.     

bFGF-mediated phosphorylation of δ-catenin increases its protein stability and the ability to induce the nuclear redistribution of β-catenin

Recently, we have shown that δ-catenin strengthened the epidermal growth factor receptor (EGFR)/Erk1/2 signaling pathway through the association between EGFR and δ-catenin. Now, we further analyzed the correlation between basic fibroblast growth factor (bFGF)/fibroblast growth factor receptor 1 (FGFR1) and δ-catenin in prostate cancer and investigated the molecular mechanism underlying the role of bFGF/FGFR1 modulation in CWR22Rv-1 (Rv-1) cells. Here, we demonstrated that bFGF phosphorylated the tyrosine residues of δ-catenin in Rv-1 cells and further proved that the bFGF mediated FGFR1/δ-catenin tyrosine phosphorylation was time dependent. Furthermore, we demonstrated that bFGF stabilized the expression of δ-catenin through weakening its association with GSK3β and enhancing its stability to induce β-catenin into the nuclear by strengthening the processing of E-cadherin. In a word, these results indicated that bFGF/FGFR1 signaling pathway could enhance the tumor progression of prostate cancer via δ-catenin.     

TM2D1 contributes the epithelial-mesenchymal transition of hepatocellular carcinoma via modulating AKT/β-catenin axis

Various epidemiology studies showed the correlation between Alzheimer’s disease (AD) and low incidence of cancer. However, the etiology underlying etiology of AD-related carcinogenesis remains largely elusive. Our study focused on characterizing the role of TM2D1 (TM2 domain containing 1) in hepatocellular carcinoma. TM2D1 is also known as β-amyloid peptide binding protein and is critical to the pathogenesis of AD. We found that TM2D1 is increasingly expressed in HCC tumors relative to the peritumoral tissues of the matched tumors and high TM2D1 expression predicts unfavorable clinical outcomes. TM2D1 overexpression induced HCC cell proliferation, migration and invasion, which was related to the epithelial-mesenchymal transition (EMT) observed in these cells. Conversely, TM2D1 depletion led to opposite phenotype in HCC. Mechanistically, we found that TM2D1 promoted Akt and β-catenin hyper-activation, which corresponded with molecular marker change in EMT signaling pathway. Taken together, our results indicated that TM2D1 played an important role in the EMT process in HCC cells by activating AKT and β-catenin signaling and may become a promising therapeutic target in HCC.     

Downregulation of long non-coding RNA MR4435-2HG suppresses breast cancer progression via the Wnt/β-catenin signaling pathway

Extensive research has contributed to the current understanding of the critical roles played by long non-coding RNAs in various types of cancer. The present study aimed to investigate the function and mechanism of the long non-coding RNA, MIR4435-2HG (also termed LINC00978), in breast cancer growth and metastasis. Using Gene Expression Profiling Interactive Analysis, an online web tool, it was revealed that MIR4435-2HG was upregulated in breast cancer tissue, and its high expression was associated with poor prognosis based on The Cancer Genome Atlas database. MIR4435-2HG knockdown increased cell apoptosis but decreased cell proliferation, migration and invasion. MIR4435-2HG knockdown increased pro-apoptotic protein expression but decreased anti-apoptotic protein expression. In addition, MIR4435-2HG knockdown leads to dysregulation of epithelial-to-mesenchymal transition-associated genes. Furthermore, knockdown of MIR4435-2HG results in inactivation of the Wnt/β-catenin signaling pathway. The results of the present study demonstrate the tumor-promoting role of MIR4435-2HG in breast cancer progression.     

Constitutive activation of integrin αvβ3 contributes to anoikis resistance of ovarian cancer cells

Epithelial ovarian cancer involves the shedding of single tumor cells or spheroids from the primary tumor into ascites, followed by their survival, and transit to the sites of metastatic colonization within the peritoneal cavity. During their flotation, anchorage‐dependent epithelial‐type tumor cells gain anoikis resistance, implicating integrins, including αvß3. In this study, we explored anoikis escape, cisplatin resistance, and prosurvival signaling as a function of the αvß3 transmembrane conformational activation state in cells suspended in ascites. A high‐affinity and constitutively signaling‐competent αvß3 variant, which harbored unclasped transmembrane domains, was found to confer delayed anoikis onset, enhanced cisplatin resistance, and reduced cell proliferation in ascites or 3D‐hydrogels, involving p27^kip upregulation. Moreover, it promoted EGF‐R expression and activation, prosurvival signaling, implicating FAK, src, and PKB/Akt. This led to the induction of the anti‐apoptotic factors Bcl‐2 and survivin suppressing caspase activation, compared to a signaling‐incapable αvß3 variant displaying firmly associated transmembrane domains. Dissecting the mechanistic players for αvß3‐dependent survival and peritoneal metastasis of ascitic ovarian cancer spheroids is of paramount importance to target their anchorage independence by reversing anoikis resistance and blocking αvß3‐triggered prosurvival signaling.

Abbreviations

CLSM

confocal laser scanning microscopy

ECM

extracellular matrix

EGF‐R

epidermal growth factor receptor

EOC

epithelial ovarian cancer

FAK

focal adhesion kinase

FIGO

Fédération Internationale de Gynécologie et d''Obstétrique

GAPDH

glyceraldehyde 3‐phosphate dehydrogenase)

GpA

glycophorin A

IMD

integrin‐mediated death

MAPK

mitogen‐activated protein kinases

PI

propidium iodide

RGD

Arg‐Gly‐Asp

TMD

transmembrane domain

     

LncRNA MEG3 inhibits retinoblastoma invasion and metastasis by inducing β-catenin degradation

In our previous study, we found that low expression of LncRNA-MEG3 was closely associated with the invasion and metastasis of retinoblastomas. The molecular mechanism by which MEG3 inactivation induces the invasion and metastasis of retinoblastoma cell lines remains unclear. We used the GEO database to analyze the expression of MEG3 in retinoblastoma tissues and MEG3-related pathways. The scratch, transwell migration, mouse tumor metastasis, and mouse fluorescence live imaging assays were performed to detect migration and invasion of retinoblastoma cell lines. The RNA pull down, electrophoretic mobility shift, RIP, co-immunoprecipitation, and ubiquitination assays were performed to analyze the molecular mechanisms. The GEO database showed that the expression of MEG3 was low in retinoblastoma tissues and was closely associated with the invasion of retinoblastoma cells and activity of the Wnt pathway. Both in vivo and in vitro experiments confirmed that MEG3 inhibited the migration and invasion of retinoblastoma cells. Cell experiments confirmed that MEG3 could promote the binding of β-catenin and GSK-3β and induce phosphorylation, ubiquitination and degradation of β-catenin indirectly. In conclusion, MEG3 can promote the degradation of β-catenin via GSK-3β, which in turn inactivates the Wnt pathway and ultimately inhibits the invasion and metastasis of retinoblastoma cells.     

Onvansertib inhibits the proliferation and improves the cisplatin-resistance of lung adenocarcinoma via β-catenin/c-Myc signaling pathway

Polo-like kinase 1 (PLK1) is a key regulator of cell division, and its abnormal expression is related to the progression and prognosis of cancers. However, the effect of PLK1 inhibitor onvansertib on the growth of lung adenocarcinoma (LUAD) has not been explored. In this study, we performed a series of bioinformatics and experimental analyses to comprehensively investigate the role of PLK1 in LUAD. We used CCK-8 assay and colony formation assay to evaluate the growth inhibitory ability of onvansertib. Furthermore, flow cytometry was applied to exploit the effects of onvansertib on cell cycle, apoptosis, and mitochondrial membrane potential. Moreover, the therapeutic potential of onvansertib was assessed in vivo by using xenograft tumor and patient-derived xenograft (PDX) models. We found that onvansertib significantly induced the apoptosis and inhibited the proliferation and migration of LUAD cells. Mechanistically, onvansertib arrested the cells at G2/M phase and enhanced the levels of reactive oxidative species in LUAD. Accordingly, onvansertib regulated the expression of glycolysis-related genes and improved the cisplatin resistance in LUAD. Notably, the protein levels of β-catenin and c-Myc were affected by onvansertib. Taken together, our findings provide insight into the function of onvansertib and shed light on the potential clinical application of onvansertib for the treatment of patients with LUAD.