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.     

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.     

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.     

Therapeutic efficacy of atezolizumab plus bevacizumab for hepatocellular carcinoma with WNT/β-catenin signal activation

Atezolizumab plus bevacizumab therapy has high response rates in patients with advanced hepatocellular carcinoma (HCC). It has been reported that HCC with immune exclusion associated with the signal activation of WNT/β-catenin is resistant to immune checkpoint inhibitors; however, to the best of our knowledge, the effectiveness of atezolizumab plus bevacizumab for HCC with WNT/β-catenin signal activation has not been reported. The present study aimed to analyze the efficacy of atezolizumab plus bevacizumab for HCC with WNT/β-catenin signal activation. A total of 24 patients who underwent liver tumor biopsy for HCC were classified into WNT/β-catenin signal activation and inactivation groups according to the expression levels of β-catenin and glutamine synthetase, which are indicative of WNT/β-catenin signal activation. The differences in the clinical responses to treatment between the groups were analyzed. A total of 15 patients had HCC with WNT/β-catenin signal activation, whereas 9 patients had HCC with WNT/β-catenin signal inactivation. There were no significant differences between both groups regarding objective responses (P=0.519) and disease control (P=0.586). In the WNT/β-catenin signal activation group, the median progression-free survival rate was 6.9 months compared with 6.2 months in the WNT/β-catenin signal inactivation group (P=0.674). Although a small number of patients was included in the present study, the present findings suggested that the efficacy of atezolizumab plus bevacizumab might be unaffected by WNT/β-catenin signal activation.     

Identification of AKT1/β-catenin mutations conferring cetuximab and chemotherapeutic drug resistance in colorectal cancer treatment

In anticancer therapy, the effectiveness of therapeutics is limited by mutations causing drug resistance. KRAS mutations are the only determinant for cetuximab resistance in patients with colorectal cancer (CRC). However, cetuximab treatment has not been fully successful in the majority of patients with wild-type (WT) KRAS. Therefore, it is important to determine new predictive mutations in CRC treatment. In the present study, the association between AKT1/β-catenin (CTNNB1) mutations with the drug resistance to cetuximab and other chemotherapeutics used in the CRC treatment was investigated by using site-directed mutagenesis, transfection, western blotting and cell proliferation inhibition assay. Cetuximab resistance was higher in the presence of AKT1 E17K, E49K and L52R mutations, as well as CTNNB1 T41A, S45F and S33P mutations compared with that of respective WT proteins. AKT1/CTNNB1 mutations were also associated with oxaliplatin, irinotecan, SN-38 and 5-fluorouracil resistance. Furthermore, mutant cell viability in oxaliplatin treatment was more effectively inhibited compared with that of the other chemotherapeutic drugs. In conclusion, AKT1/CTNNB1 mutations may be used as an important predictive biomarker in CRC treatment.     

CDH6‐activated αIIbβ3 crosstalks with α2β1 to trigger cellular adhesion and invasion in metastatic ovarian and renal cancers

Cadherin 6 (CDH6) is significantly overexpressed in advanced ovarian and renal cancers. However, the role of CDH6 in cancer metastasis is largely unclear. Here, we investigated the impact of CDH6 expression on integrin‐mediated metastatic progression. CDH6 preferentially bound to αIIbβ3 integrin, a platelet receptor scarcely expressed in cancer cells, and this interaction was mediated through the cadherin Arginine–glycine–aspartic acid (RGD) motif. Furthermore, CDH6 and CDH17 were found to interact with α2β1 in αIIbβ3^low cells. Transient silencing of CDH6, ITGA2B, or ITGB3 genes caused a significant loss of proliferation, adhesion, invasion, and lung colonization through the downregulation of SRC, FAK, AKT, and ERK signaling. In ovarian and renal cancer cells, integrin αIIbβ3 activation appears to be a prerequisite for proper α2β1 activation. Interaction of αIIbβ3 with CDH6, and subsequent αIIbβ3 activation, promoted activation of α2β1 and cell adhesion in ovarian and renal cancer cells. Additionally, monoclonal antibodies specific to the cadherin RGD motif and clinically approved αIIbβ3 inhibitors could block pro‐metastatic activity in ovarian and renal tumors. In summary, the interaction between CDH6 and αIIbβ3 regulates α2β1‐mediated adhesion and invasion of ovarian and renal cancer metastatic cells and constitutes a therapeutic target of broad potential for treating metastatic progression.     

The Wnt/β-catenin signaling pathway in the tumor microenvironment of hepatocellular carcinoma

The Wnt/β-catenin signaling pathway regulates many aspects of tumor biology, and many studies have focused on the role of this signaling pathway in tumor cells. However, it is now clear that tumor development and metastasis depend on the two-way interaction between cancer cells and their environment, thereby forming a tumor microenvironment (TME). In this review, we discuss how Wnt/β-catenin signaling regulates cross-interactions among different components of the TME, including immune cells, stem cells, tumor vasculature, and noncellular components of the TME in hepatocellular carcinoma. We also investigate their preclinical and clinical insights for primary liver cancer intervention, and explore the significance of using Wnt/β-catenin mutations as a biomarker to predict resistance in immunotherapy.     

Chromobox 4 facilitates tumorigenesis of lung adenocarcinoma through the Wnt/β-catenin pathway

Chromobox 4 (CBX4) is a core component of polycomb-repressive complex 1 with important roles in cancer biology and tissue homeostasis. Aberrant expression of CBX4 has been implicated in several human malignancies. However, its role and underlying mechanisms in the tumorigenesis of lung adenocarcinoma (LUAD) have not been defined in vivo. Here, we found that expression of CBX4 was frequently up-regulated in human LUAD samples and correlated with poor patient survival. Importantly, genetic ablation of CBX4 greatly dampened lung tumor formation and improved survival in the Kras^G12D/P53^L/L (KP) autochthonous mouse model of LUAD. In addition, CBX4 depletion significantly inhibited proliferation and anchorage-independent growth of KP mouse embryonic fibroblasts. Moreover, ectopic CBX4 expression clearly promoted proliferation and anchorage-independent growth in both human and mouse LUAD cells, whereas silencing of CBX4 exerted opposite effects. Mechanistically, CBX4 promoted growth of LUAD cells through activation of the Wnt/β-catenin pathway. Furthermore, expression levels of CBX4 were positively correlated with β-catenin in human LUAD samples. In conclusion, our data suggest that CBX4 plays an oncogenic role via the Wnt/β-catenin pathway and could serve as a potential therapeutic target in LUAD.     

RNF43 overexpression attenuates the Wnt/β-catenin signalling pathway to suppress tumour progression in cholangiocarcinoma

RING finger protein 43 (RNF43) is a ubiquitin E3 ligase that negatively regulates Wnt/β-catenin signalling. Mutation, inactivation and downregulation of RNF43 in cholangiocarcinoma (CCA) are associated with a less favourable prognosis. Since the functional role of RNF43 in CCA has not yet been demonstrated, the present study aimed to assess the effect of its overexpression in mediating CCA suppression via Wnt/β-catenin signalling pathway inhibition. Accordingly, RNF43 was overexpressed, and various malignant phenotypic changes studied, including cell proliferation, cell migration, chemotherapeutic sensitivity and the expression of several Wnt/β-catenin target genes. Overexpression of RNF43 in the CCA cell-line KKU-213B hindered activation of Wnt/β-catenin signalling, evidenced by: i) Accumulation of β-catenin in the cytoplasmic fraction and downregulation of several known Wnt target genes at the mRNA level [AXIN2, survivin (BIRC5), CCND1, MMP-7, c-MYC and ABCB1 (MDR1)]; ii) a reduction of cell proliferation; iii) a significant decrease in KKU-213B cell migration with RNF43 overexpression via upregulation of E-cadherin (CDH1); and iv) a reduction in N-cadherin (CDH2), MMP-2, MMP-7 and MMP-9. In addition, overexpression of RNF43 increased 5-fluorouracil sensitivity and downregulation of ABC transporter genes [including ABCB1 and ABCC1 (MRP1)]. The current results demonstrate a functional role for RNF43 in CCA by: i) Blocking β-catenin nuclear translocation; and ii) the subsequent downregulation of Wnt/β-catenin target genes (the latter being involved in the progression of CCA and chemotherapeutic drug susceptibility). Therefore, the present findings suggest that RNF43 could serve a tumour suppressive role in CCA.     

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.     

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.     

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

     

Perphenazine and prochlorperazine decrease glioblastoma U-87 MG cell migration and invasion: Analysis of the ABCB1 and ABCG2 transporters,E-cadherin, α-tubulin and integrins (α3, α5, and β1) levels

Glioblastoma multiforme is the most frequent type of malignant brain tumor, and is one of the most lethal and untreatable human tumors with a very poor survival rate. Therefore, novel and effective strategies of treatment are required. Integrins play a crucial role in the regulation of cellular adhesion and invasion. Integrins and α-tubulin are very important in cell migration, whereas E-cadherin plays a main role in tumor metastasis. Notably, drugs serve a crucial role in glioblastoma treatment; however, they have to penetrate the blood-brain barrier (BBB) to be effective. ABC transporters, including ATP binding cassette subfamily B member 1 (ABCB1) and ATP binding cassette subfamily G member 2 (ABCG2), are localized in the brain endothelial capillaries of the BBB, have a crucial role in the development of multidrug resistance and are modulated by phenothiazine derivatives. The impact of perphenazine and prochlorperazine on the motility of human Uppsala 87 malignant glioma (U87-MG) cells was evaluated using a wound-healing assay, cellular migration and invasion were assessed by Transwell assay, and the protein expression levels of ABCB1, ABCG2, E-cadherin, α-tubulin and integrins were determined by western blotting. The present study explored the effects of perphenazine and prochlorperazine on the levels of ABCB1, ABCG2, E-cadherin, α-tubulin and integrins (α3, α5, and β1), as well as on the migratory and invasive ability of U87-MG cells. The results suggested that perphenazine and prochlorperazine may modulate the expression levels of multidrug resistance proteins (they decreased ABCB1 and increased ABCG2 expression), E-cadherin, α-tubulin and integrins, and could impair the migration and invasion of U-87 MG cells. In conclusion, the decrease in migratory and invasive ability following treatment with phenothiazine derivatives due to the increase in ABCG2 and E-cadherin expression, and decrease in α-tubulin and integrins expression, may suggest that research on perphenazine and prochlorperazine in the treatment of glioblastoma is worth continuing.     

SIK2 represses AKT/GSK3β/β‐catenin signaling and suppresses gastric cancer by inhibiting autophagic degradation of protein phosphatases

Salt‐inducible kinase 2 (SIK2) is an important regulator in various intracellular signaling pathways related to apoptosis, tumorigenesis and metastasis. However, the involvement of SIK2 in gastric tumorigenesis and the functional linkage with gastric cancer (GC) progression remain to be defined. Here, we report that SIK2 was significantly downregulated in human GC tissues, and reduced SIK2 expression was associated with poor prognosis of patients. Overexpression of SIK2 suppressed the migration and invasion of GC cells, whereas knockdown of SIK2 enhanced cell migratory and invasive capability as well as metastatic potential. These changes in the malignant phenotype resulted from the ability of SIK2 to suppress epithelial–mesenchymal transition via inhibition of AKT/GSK3β/β‐catenin signaling. The inhibitory effect of SIK2 on AKT/GSK3β/β‐catenin signaling was mediated primarily through inactivation of AKT, due to its enhanced dephosphorylation by the upregulated protein phosphatases PHLPP2 and PP2A. The upregulation of PHLPP2 and PP2A was attributable to SIK2 phosphorylation and activation of mTORC1, which inhibited autophagic degradation of these two phosphatases. These results suggest that SIK2 acts as a tumor suppressor in GC and may serve as a novel prognostic biomarker and therapeutic target for this tumor.

Abbreviations

AMPK

AMP‐activated protein kinase

Co‐IP

co‐immunoprecipitation

EMT

epithelial–mesenchymal transition

GAPDH

glyceraldehyde‐3‐phosphate dehydrogenase

GC

gastric cancer

GEO

Gene Expression Omnibus

H&E

hematoxylin and eosin

IHC

immunohistochemistry

mTOR

mechanistic target of rapamycin

NC

negative control

PHLPP

PH domain leucine‐rich repeat protein phosphatase

PP2A

protein phosphatase 2A

qRT‐PCR

quantitative real‐time polymerase chain reaction

SIK2

salt‐inducible kinase 2

TCF/LEF

T cell factor/lymphoid enhancer‐binding factor

TCGA

The Cancer Genome Atlas