Coactivation of NF-κB and Notch signaling is sufficient to induce B-cell transformation and enables B-myeloid conversion.

NF-κB and Notch signaling can be simultaneously activated in a variety of B-cell lymphomas. Patients with B-cell lymphoma occasionally develop clonally related myeloid tumors with poor prognosis. Whether concurrent activation of both pathways is sufficient to induce B-cell transformation and whether the signaling initiates B-myeloid conversion in a pathological context are largely unknown. Here, we provide genetic evidence that concurrent activation of NF-κB and Notch signaling in committed B cells is sufficient to induce B-cell lymphomatous transformation and primes common progenitor cells to convert to myeloid lineage through dedifferentiation, not transdifferentiation. Intriguingly, the converted myeloid cells can further transform, albeit at low frequency, into myeloid leukemia. Mechanistically, coactivation of NF-κB and Notch signaling endows committed B cells with the ability to self renew. Downregulation of BACH2, a lymphoma and myeloid gene suppressor, but not upregulation of CEBPα and/or downregulation of B-cell transcription factors, is an early event in both B-cell transformation and myeloid conversion. Interestingly, a DNA hypomethylating drug not only effectively eliminated the converted myeloid leukemia cells, but also restored the expression of green fluorescent protein, which had been lost in converted myeloid leukemia cells. Collectively, our results suggest that targeting NF-κB and Notch signaling will not only improve lymphoma treatment, but also prevent the lymphoma-to-myeloid tumor conversion. Importantly, DNA hypomethylating drugs might efficiently treat these converted myeloid neoplasms.

RASSF10 suppresses lung cancer proliferation and invasion by decreasing the level of phosphorylated LRP6.

Ras-association domain family (RASSF) proteins exert distinct cellular functions. The expression of RASSF10 in non-small cell lung cancer and its underlying mechanism have not been reported. Herein, we explored the roles of RASSF10 in lung cancer cells and potential molecular mechanisms. We found low RASSF10 expression in lung cancer specimens, which was associated with low differentiation, advanced pTNM stage, positive lymph node metastasis, and poor prognosis in patients. Furthermore, RASSF10 overexpression inhibited the proliferation and invasion of lung cancer cells, which was the result of Wnt signaling suppression. However, we found that RASSF10 had no influence on Hippo signaling, while RASSF10 bound to LRP6 via the coiled-coil domains and reduced p-LRP6 level, eventually prohibiting ß-catenin nuclear translocation. However, deleting the coiled-coil domains ablated this function. These findings expound the interaction between RASSF10 and LRP6 and uncover a potential link between N-terminal RASSFs and the Wnt pathway.

Lipopolysaccharide stimulates endogenous ß-glucuronidase via PKC/NF-κB/c-myc signaling cascade: a possible factor in hepatolithiasis formation.

Hepatolithiasis is commonly encountered in Southeastern and Eastern Asian countries, but the pathogenesis mechanism of stone formation is still not well understood. Now, the role of endogenous ß-glucuronidase in pigment stones formation is being gradually recognized. In this study, the mechanism of increased expression and secretion of endogenous ß-glucuronidase during hepatolithiasis formation was investigated. We assessed the endogenous ß-glucuronidase, c-myc, p-p65, and p-PKC expression in liver specimens with hepatolithiasis by immunohistochemical staining, and found that compared with that in normal liver samples, the expression of endogenous ß-glucuronidase, c-myc, p-p65, and p-PKC in liver specimens with hepatolithiasis significantly increased, and their expressions were positively correlated with each other. Lipopolysaccharide (LPS) induced increased expression of endogenous ß-glucuronidase and c-myc in hepatocytes and intrahepatic biliary epithelial cells in a dose- and time-dependent manner, and endogenous ß-glucuronidase secretion increased, correspondingly. C-myc siRNA transfection effectively inhibited the LPS-induced expression of endogenous ß-glucuronidase. Furthermore, NF-κB inhibitor pyrrolidine dithiocarbamate or PKC inhibitor chelerythrine could effectively inhibit the LPS-induced expression of c-myc and endogenous ß-glucuronidase, and the expression of p-p65 was also partly inhibited by chelerythrine. Our clinical observations and experimental data indicate that LPS could induce the increased expression and secretion of endogenous ß-glucuronidase via a signaling cascade of PKC/NF-κB/c-myc in hepatocytes and intrahepatic biliary epithelial cells, and endogenous ß-glucuronidase might play a possible role in the formation of hepatolithiasis.

Rab23 is overexpressed in human astrocytoma and promotes cell migration and invasion through regulation of Rac1.

Rab23 overexpression has been implicated in several human cancers. However, its biological roles and molecular mechanism in astrocytoma have not been elucidated. The aim of this study is to explore clinical significance and biological roles of Rab23 in astrocytoma. We observed negative Rab23 staining in normal astrocytes and positive staining in 39 out of 86 (45 %) astrocytoma specimens using immunohistochemistry. The positive rate of Rab23 was higher in grades III and IV (56.5 %, 26/46) than grades I + II astrocytomas (32.5 %, 13/40, p < 0.05). Transfection of Rab23 plasmid was performed induced A172 cell proliferation, colony formation, invasion, and migration, while Rab23 depletion with siRNA reduced these abilities of U87 cells. In addition, we found that Rab23 transfection upregulated while its depletion reduced Rac1 activity. Treatment of transfected cells with a Rac1 inhibitor decreased Rac1 activity and invasion. In conclusion, Rab23 serves as an important oncoprotein in human astrocytoma by regulating cell invasion and migration through Rac1 activity.

RASSF4 is downregulated in nonsmall cell lung cancer and inhibits cancer cell proliferation and invasion.

RASSF4 has been implicated as a tumor suppressor in several human cancers. Its clinical significance and biological characteristics in human nonsmall cell lung cancer (NSCLC) have not been explored yet. In this study, we explored expression pattern of RASSF4 in 89 NSCLC specimens. The results showed that RASSF4 was downregulated in 36/89 NSCLC tissues compared with normal tissue. RASSF4 downregulation significantly associated with advanced TNM stage, positive nodal status, and poor prognosis. We examined RASSF4 protein expression in normal lung epithelial cell line and lung cancer lines. We found that RASSF4 expression was downregulated in four of seven lung cancer cell lines compared with normal bronchial epithelial cells. RASSF4 plasmid transfection was performed in H460 and A549 cell lines. RASSF4 overexpression inhibited proliferation, colony formation, and invading ability. In addition, we identified that RASSF4 could inhibit cell cycle progression with downregulation of cyclin D1. Expression of invasion-related protein MMP2, MMP9 was also decreased. In conclusion, the present study suggested that RASSF4 serves as an important tumor suppressor in NSCLC.

Overexpression of CASS4 promotes invasion in non-small cell lung cancer by activating the AKT signaling pathway and inhibiting E-cadherin expression.

The role of Crk-associated substrate (CAS) family members in regulating invasion and metastasis has been described in several cancers. As the fourth member of the CAS family, CASS4 is also related with positive lymph node metastasis and poor prognosis in lung cancer. However, the underlying mechanisms and downstream effectors of CASS4 in the development and progression of non-small cell lung cancer (NSCLC) remain unclear. In this study, CASS4 overexpression inhibited E-cadherin expression and enhanced invasion in NSCLC cell line transfected with CASS4 plasmid, while CASS4 depletion upregulated E-cadherin expression and inhibited invasion in NSCLC cell line transfected with CASS4 siRNA. The effect of CASS4 overexpression in facilitating invasion of NSCLC cells was reversed by restoring E-cadherin expression, which indicates that CASS4 may promote invasion by inhibiting E-cadherin expression. Subsequent immunohistochemistry results confirmed that CASS4 overexpression correlated with loss of E-cadherin expression. We next investigated the phosphorylation levels of focal adhesion kinase (FAK), p38, extracellular signal-related kinase (ERK), and AKT after CASS4 plasmid or CASS4 siRNA transfection. CASS4 facilitated AKT (Ser473) phosphorylation. Treatment with an AKT phosphorylation inhibitor reversed the increased invasive capacity and downregulation of E-cadherin protein induced by CASS4 overexpression. Taken together, the present results indicate that CASS4 may promote NSCLC invasion by activating the AKT signaling pathway, thereby inhibiting E-cadherin expression.

Dishevelled-3 activates p65 to upregulate p120-catenin transcription via a p38-dependent pathway in non-small cell lung cancer.

Dishevelled-3 (Dvl-3) and p120-catenin (p120ctn) have abnormal expression in non-small cell lung cancer (NSCLC), which is associated with poor prognosis. Dvl-3 upregulates p120ctn transcription in NSCLC cells, but the mechanism is unknown. Here we transiently transfected Dvl-3 cDNA to NSCLC cells. Dvl-3 transfection is sufficient for induction of p38 signaling. In turn, Dvl-3 induces p38-mediated activation of the p65 so as to facilitate its nuclear translocation. Treatment with SB203580 (p38 inhibitor) or BAY 11-7082 (IκB-α phosphorylation inhibitor) suppresses Dvl-3 induced activation of p65. The results further show that active p65 interacts with PAX2 promoter to increase the expression of PAX2 and then PAX2 binds to p120ctn promoter so as to upregulate p120ctn gene transcription. Moreover, Dvl-3 transfection enhanced the binding of active p65 to Sp1 so as to decrease the binding of Sp1 to p120ctn promoter. The above-mentioned effects are linked to biological behavior of non-small cell lung cancer cells. These findings confirm that p38 and PAX2 are important for the Dvl-3 induced upregulation of p120ctn. Dvl-3 activates a p38 → p65 → PAX2 → p120ctn pathway to affect biological behavior of NSCLC cells.

CRKL protein overexpression enhances cell proliferation and invasion in pancreatic cancer.

CRKL is an adapter protein which is overexpressed in many malignant tumors and plays crucial roles in tumor progression. However, expression pattern and biological roles of CRKL in pancreatic cancer have not been examined. In the present study, we found that CRKL expression in pancreatic cancer specimens was higher than that in normal pancreatic tissues. Colony formation assay and Matrigel invasion assay showed that the overexpression of CRKL in Bxpc3 and Capan2 cell lines with low endogenous expression increased cell proliferation and invasion. Flow cytometry showed that CRKL promoted cell proliferation by facilitating cell cycle. Further analysis of cell cycle- and invasion-related molecules showed that CRKL upregulated cyclin D1, cyclin A, matrix metalloproteinase 2 (MMP2) expression, and phosphorylated extracellular signal (ERK)-regulated kinase. In conclusion, our study demonstrated that CRKL was overexpressed in human pancreatic cancers and contributed to pancreatic cancer cell proliferation and invasion through ERK signaling.

Derlin-1 is overexpressed in non-small cell lung cancer and promotes cancer cell invasion via EGFR-ERK-mediated up-regulation of MMP-2 and MMP-9.

Previous studies indicated a role of Derlin-1 in human cancers; however, its expression pattern in non-small cell lung cancer (NSCLC) and the molecular mechanism of Derlin-1 on cancer progression have not been characterized. In the present study, Derlin-1 expression was examined in lung cancer cell lines and human tissues. Derlin-1 overexpression correlated with pTNM stage, lymph node metastasis, and poor overall survival. siRNA knockdown of Derlin-1 impaired anchorage-dependent and anchorage-independent cell growth and invasion in A549 and H1299 cell lines, and its overexpression promoted proliferation and invasion in HBE and LTE cell lines. Derlin-1 depletion decreased matrix metalloproteinase (MMP)-2/9 at both protein and mRNA levels, with decreased MAP kinase/extracellular signal-regulated kinase (ERK)/ERK phosphorylation. Derlin-1 overexpression up-regulated MMP-2/9 expression and ERK phosphorylation, which could be reversed by MAP kinase/ERK kinase inhibitor, PD98059. The effect of Derlin-1 on MMP-2/9 up-regulation was abolished in ERK1/2 siRNA-treated cells. Further analysis showed that Derlin-1 overexpression induced EGFR phosphorylation. EGFR inhibitor blocked Derlin-1-mediated up-regulation of EGFR and ERK phosphorylation. MMP-2/9 and p-ERK up-regulation by Derlin-1 was partly blocked in EGFR-depleted cells with siRNA treatment. Immunoprecipitation confirmed the association between Derlin-1 and EGFR. In summary, our results showed that Derlin-1 is overexpressed in NSCLC and promotes invasion by EGFR-ERK-mediated up-regulation of MMP-2 and MMP-9. Derlin-1 may serve as a therapeutic target for NSCLC.

Diversin is overexpressed in human gliomas and its depletion inhibits proliferation and invasion.

Previous study indicated diversin overexpression in human cancers. However, its expression pattern in human gliomas and the molecular mechanisms of diversin on cancer progression have not been characterized. In the present study, diversin expression was investigated in 105 glioma specimens using immunohistochemistry. Negative staining was observed in normal glial cells, and positive staining was found in 33 out of 105 (31.4 %) glioma specimens. Diversin overexpression correlated with advanced tumor grade (p < 0.05). Small interfering RNA (siRNA) knockdown was performed in U87 and TG905 cell lines with high endogenous diversin expression. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation assay showed that diversin knockdown inhibited glioma cell growth. Matrigel invasion assay showed that diversin depletion inhibited cell invasion. In addition, messenger RNA (mRNA) and protein levels of matrix metallopeptidase 9 (MMP9) were downregulated after siRNA treatment. In conclusion, diversin is overexpressed in human glioma and regulates glioma cell proliferation and invasion, possibly through MMP9.

Examining Nek2 as a better proliferation marker in non-small cell lung cancer prognosis.

The purpose of this study is to identify a better potential biomarker for the prognosis of patients with non-small cell lung cancer (NSCLC). The expressions of Nek2, MCM7, and Ki-67 were evaluated in 270 NSCLC tissues using immunohistochemical and immunofluorescence techniques. Associations between protein expression and clinical pathologic characters were assessed, and the impact on overall survival was analyzed. We detected high levels of Nek2, MCM7, and Ki-67 expression in 25.9, 35.2, and 24.4 % of NSCLC tissues, respectively. Overexpressions of Nek2 were detected more frequently in high T-stage and N-stage cases (P = 0.000, 0.011). The expressions of Nek2, MCM7, and Ki-67 were correlated with each other. Kaplan-Meier curves indicated that patients with overexpression of Nek2, MCM7, and Ki-67 had a poorer overall survival rate compared to those with low expression for all stages (P = 0.000). In particular, the patients with Nek2 overexpression had the most negative prognosis. Multivariate Cox regression analysis showed that Nek2, MCM7, and Ki-67 are independent prognostic indicators for NSCLC. Our data suggest that among Nek2 kinase, MCM7, and Ki-67, it is Nek2 kinase that is the more effective tumor proliferation marker of poor prognosis for NSCLC patients. Thus, Nek2 may represent a new potential target for NSCLC therapeutic intervention.

Philadelphia chromosome-positive T-cell acute lymphoblastic leukemia: a case report.

Philadelphia chromosome-positive (Ph+) T-cell acute lymphoblastic leukemia (T-ALL) is a rare and aggressive type of acute leukemia. The Philadelphia chromosome is the hallmark of chronic myeloid leukemia (CML). The differentiation between Ph+ T-ALL and T-cell lymphoblastic crisis of CML may be problematic in some cases. Here, we report a rare case of de novo Ph+ T-ALL that presented a diagnostic challenge. The overall clinical, immunophenotypic, cytogenetic, and xenotransplantation results suggest a diagnosis of Ph+ T-ALL. The patient was treated with induction chemotherapy including imatinib followed by haploidentical stem cell transplantation and achieved complete remission.

Overexpression of CRKL correlates with poor prognosis and cell proliferation in non-small cell lung cancer.

Crk-Like (CRKL) is an adapter protein that has crucial roles in multiple biological processes, including cell proliferation, adhesion, and migration. Amplification of CRKL gene was found in non-small cell lung cancer (NSCLC). However, the expression pattern of CRKL protein and its clinical significance in human NSCLC have not been well characterized to date. In this study, expression of CRKL was evaluated in 131 NSCLC tissues by immumohistochemistry. CRKL protein was up-regulated in the lung carcinomas compared with adjacent normal lung tissue. Overexpression of CRKL was found in 58 of 131 (44.3%) NSCLC samples and correlated with poor tumor differentiation (P = 0.0042), histological type (adenocarcinoma; P = 0.001), advanced p-TNM stage (P = 0.0004), nodal metastasis (P = 0.0273), high proliferation index (P = 0.0062) and poor overall survival (P = 0.0084). Further univariate and multivariate analysis showed a significant association of CRKL overexpression and worse overall survival in lung cancer patients. In addition, overexpression of CRKL in HBE and H1299 cell lines promoted cell proliferation by facilitating cell cycle progression. Further analysis of cell cycle related molecules showed that CRKL induced cyclin D1, cyclin B1 expression, and increased Rb phosphorylation. In conclusion, this study demonstrated overexpression of CRKL correlated with poor prognosis and lung cancer proliferation by cell cycle regulation.

Ataxia-telangiectasia group D complementing gene (ATDC) upregulates matrix metalloproteinase 9 (MMP-9) to promote lung cancer cell invasion by activating ERK and JNK pathways.

Although the expression pattern and biological functions of ataxia-telangiectasia group D complementing gene (ATDC) had been implicated in several types of cancer, the roles and potential mechanisms of ATDC in lung cancer cell invasion are still ambiguous. In this study, we used gain- and loss-of-function analyses to explore the roles and potential mechanisms of ATDC in lung cancer cell invasion. siRNA knockdown of ATDC impaired cell invasion in A549 and H1299 cell lines, and its overexpression promoted cell invasion in HBE cell line. ATDC may contribute to the invasive ability of lung cancer cells by promoting the expression of invasion-related matrix metalloproteinase 9 (MMP-9). In addition, ATDC increased activating protein 1 (AP-1) reporter luciferase activity and the protein and mRNA levels of c-Jun and c-Fos. We further demonstrated that the roles of ATDC on cell invasion, MMP-9 upregulation, and AP-1 activation were dependent on extracellular signal-regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK) pathway activation, and ERK inhibitor U0126 or JNK inhibitor SP600125 blocked these effects of ATDC. These results suggested that ATDC upregulates MMP-9 to promote lung cancer cell invasion by activating ERK and JNK pathways.

Abnormal expression of Pygopus 2 correlates with a malignant phenotype in human lung cancer.

BACKGROUND: Pygopus 2 (Pygo2) is a Pygo family member and an important component of the Wnt signaling transcriptional complex. Despite this data, no clinical studies investigating Pygo2 expression in lung cancer have yet been reported. METHODS: In the present study, the expression patterns of Pygo2 were evaluated by immunochemistry in 168 patients with non-small cell lung cancer (NSCLC). We used small interfering RNA (siRNA) to specifically silence Pygo2, and investigated its effect on cell growth by an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis in human lung cancer cell lines. RESULTS: Immunohistochemical analysis showed low expression of Pygo2 in normal lung tissues and increased nuclear expression in lung cancer tissues, either with or without perinuclear expression. Abnormal Pygo2 expression was associated with poor differentiation and a high Tumor (T), Node (N) and Metastases (M) stage in NSCLC patients, and correlated with poor prognosis. Using MTT assay we observed that Pygo2 downregulation inhibited cell proliferation; in addition, flow cytometry analysis showed that Pygo2 knockdown induced apoptosis and increased numbers of G1-phase cells and a reduction in S-phase cells. CONCLUSIONS: We therefore conclude that abnormal Pygo2 protein expression may be a marker for advanced NSCLC. Furthermore, Pygo2 knockdown suppresses cell growth.

Rsf-1 is overexpressed in non-small cell lung cancers and regulates cyclinD1 expression and ERK activity.

Rsf-1 (HBXAP) was recently reported to be overexpressed in various cancers and associated with the malignant behavior of cancer cells. However, the expression of Rsf-1 in primary lung cancer and its biological roles in non-small cell lung cancer (NSCLC) have not been reported. The molecular mechanism of Rsf-1 in cancer aggressiveness remains ambiguous. In the present study, we analyzed the expression pattern of Rsf-1 in NSCLC tissues and found that Rsf-1 was overexpressed at both the mRNA and protein levels. There was a significant association between Rsf-1 overexpression and TNM stage (p=0.0220) and poor differentiation (p=0.0013). Furthermore, knockdown of Rsf-1 expression in H1299 and H460 cells with high endogenous Rsf-1 expression resulted in a decrease of colony formation ability and inhibition of cell cycle progression. Rsf-1 knockdown also induced apoptosis in these cell lines. Further analysis showed that Rsf-1 knockdown decreased cyclin D1 expression and phospho-ERK levels. In conclusion, Rsf-1 is overexpressed in NSCLC and contributes to malignant cell growth by cyclin D1 and ERK modulation, which makes Rsf-1 a candidate therapeutic target in lung cancer.

Downregulation of Mig-6 in nonsmall-cell lung cancer is associated with EGFR signaling.

Downregulation of Mig-6 expression has been implicated in several human cancers and its loss can lead to prolonged activation of EGFR and carcinogenesis. The present study aimed to investigate the clinical significance of loss of Mig-6 expression in nonsmall-cell lung cancer (NSCLC) and the biological functions of Mig-6 in NSCLC cell lines. Mig-6 expression was downregulated in 47/91 (51.6%) cases of NSCLC that were examined. Mig-6 downregulation significantly correlated with poor differentiation (P = 0.0131), histological type (P = 0.0021), and EGFR expression (P = 0.003). In addition, knockdown of Mig-6 expression in H1299 and BE1 cells promoted EGF-induced tumor cell proliferation and migration. Furthermore, Mig-6 knockdown led to a significant increase in phospho-AKT, phospho-ERK, phospho-EGFR as well as MMP-2 and MMP-9 levels. These results indicate that downregulated Mig-6 in NSCLC tissues may serve as a new marker that can predict the activation of EGFR signaling pathway.

Rsf-1 overexpression correlates with poor prognosis and cell proliferation in colon cancer.

Rsf-1 (HBXAP) was recently reported to be overexpressed in various cancers and associated with the malignant behavior of cancer cells. However, the expression of Rsf-1 and its biological roles in colon cancer have not been reported. The molecular mechanism of Rsf-1 in cancer aggressiveness remains ambiguous. In the present study, we analyzed the expression pattern of Rsf-1 in colon cancer tissues and found that Rsf-1 was overexpressed in 50.4 % of colon cancer specimens. There was a significant association between Rsf-1 overexpression and TNM stage (p = 0.0205), lymph node metastasis (p = 0.0025), and poor differentiation (p = 0.0235). Furthermore, Rsf-1 overexpression correlated with a poor prognosis in colon cancer patients (p = 0.0011). In addition, knockdown of Rsf-1 expression in HT29 and HCT116 cells with high endogenous Rsf-1 expression decrease cell proliferation and colony formation ability. Further analysis showed that Rsf-1 knockdown decreased cyclin E expression and phospho-Rb level. In conclusion, Rsf-1 is overexpressed in colon cancers and contributes to malignant cell growth by cyclin E and phospho-Rb modulation, which makes Rsf-1 a candidate therapeutic target in colon cancer.