SHP-1 Expression Research Articles

Significance and expression of Shp2 in liver cancer stem cell

Objective To study the expression of Shp2 in liver cancer stem cells and the significance of liver stem cells regulation,and then clarify the molecular mechanism of Shp2 in regulating the liver cancer stem cells,explore its feasibility as a therapeutic target for liver cancer and liver cancer biomarkers for individualized treatment. Methods Real-time PCR was used to detect the expression of Shp2 in chemotherapy resistant in HCC cells xenografts.In addition,the expression of Shp2 was detected by real-time PCR technique in stem cell enriched HCC cells using a variety of sorting techniques.The interfering Shp2 cell line was used to define the possible regulation mechanism of Shp2 on liver cancer stem cells by Western blot method. Results In vivo experiments showed that Shp2 expression increased in chemotherapy resistant liver cancer xenografts.What′s more,in vitro experiments,the expression of Shp2 was elevated in the liver cancer stem cells.The expression of Shp2 is closely related to the b-catenin in the interfering Shp2 HCC cell lines,suggesting that Shp2 may promote the formation of liver cancer stem cells through the Wnt/b-catenin pathway. Conclusion In this study,we found that the activation of Wnt/b-catenin signaling pathway by Shp2 in HCC cells promotes cancer stem cell expansion,which suggesting that Shp2 maybe as a biomarker of liver cancer stem cells,and provide a new molecular target and clinical basis for clinical transformation study. Key words: Hepatocellular carcinoma; Shp2; Liver cancer stem cells; Chemotherapy resistance

SHP2 overexpression enhances the invasion and metastasis of ovarian cancer in vitro and in vivo.

PurposeSHP2 has roles in a variety of signal transduction pathways and in many important cellular processes, including proliferation, differentiation, movement regulation, and apoptosis. In addition, SHP2 expression is closely associated with multiple types of malignancies. In this study, we examined the role of SHP2 in epithelial ovarian cancer.Patients and methodsSHP2 expression in cancer and normal ovarian tissue specimens was evaluated by immunohistochemical staining and Western blot analyses. The correlation between the SHP2 expression level and clinicopathological features was analyzed. The role of SHP2 in epithelial ovarian cancer was evaluated by assessing SHP2 expression patterns in vitro and in vivo, and activation of the PI3K/AKT pathway was examined.ResultsSHP2 is expressed at higher levels in ovarian cancer tissues than in normal ovarian tissues and in an ovarian cancer cell line than in a normal ovarian cell line. On the basis of these findings, SHP2 is overexpressed in ovarian cancer both in vitro and in vivo. In addition, SHP2 overexpression is associated with tumor stage and differentiation, enhanced cell proliferation and invasion, and tumorigenesis and metastasis.ConclusionSHP2 overexpression enhances ovarian tumor proliferation and invasion by activating the PI3K-AKT axis, indicating that SHP2 potentially plays a direct role in the pathogenesis of ovarian epithelial cell cancer. These novel findings provide key insights that are applicable to basic cancer research and to the prevention and treatment of cancer.

Reptin regulates insulin-stimulated Akt phosphorylation in hepatocellular carcinoma via the regulation of SHP-1/PTPN6.

Hepatocellular carcinoma (HCC) is the main primary cancer of the liver. Many studies have shown that insulin resistance is a risk factor for HCC. We previously discovered the overexpression and oncogenic role of the Reptin/RUVBL2 ATPase in HCC. Here, we found that Reptin silencing enhanced insulin sensitivity in 2 HCC cell lines, as shown by a large potentiation of insulin-induced AKT phosphorylation on Ser473 and Thr308, and of downstream signalling. Reptin silencing did not affect the tyrosine phosphorylation of the insulin receptor nor of IRS1, but it enhanced the tyrosine phosphorylation of the p85 subunit of PI3K. The expression of the SHP-1/PTPN6 phosphatase, which dephosphorylates p85, was reduced after Reptin depletion. Forced expression of SHP-1 restored a normal AKT phosphorylation after insulin treatment in cells where Reptin was silenced, demonstrating that the downregulation of SHP1 is mechanistically linked to increased Akt phosphorylation. In conclusion, we have uncovered a new function for Reptin in regulating insulin signalling in HCC cells via the regulation of SHP-1 expression. We suggest that the regulation of insulin sensitivity by Reptin contributes to its oncogenic action in the liver.

The tyrosine kinase inhibitor nintedanib activates SHP-1 and induces apoptosis in triple-negative breast cancer cells

Triple-negative breast cancer (TNBC) remains difficult to treat and urgently needs new therapeutic options. Nintedanib, a multikinase inhibitor, has exhibited efficacy in early clinical trials for HER2-negative breast cancer. In this study, we examined a new molecular mechanism of nintedanib in TNBC. The results demonstrated that nintedanib enhanced TNBC cell apoptosis, which was accompanied by a reduction of p-STAT3 and its downstream proteins. STAT3 overexpression suppressed nintedanib-mediated apoptosis and further increased the activity of purified SHP-1 protein. Moreover, treatment with either a specific inhibitor of SHP-1 or SHP-1-targeted siRNA reduced the apoptotic effects of nintedanib, which validates the role of SHP-1 in nintedanib-mediated apoptosis. Furthermore, nintedanib-induced apoptosis was attenuated in TNBC cells expressing SHP-1 mutants with constantly open conformations, suggesting that the autoinhibitory mechanism of SHP-1 attenuated the effects of nintedanib. Importantly, nintedanib significantly inhibited tumor growth via the SHP-1/p-STAT3 pathway. Clinically, SHP-1 levels were downregulated, whereas p-STAT3 was upregulated in tumor tissues, and SHP-1 transcripts were associated with improved disease-free survival in TNBC patients. Our findings revealed that nintedanib induces TNBC apoptosis by acting as a SHP-1 agonist, suggesting that targeting STAT3 by enhancing SHP-1 expression could be a viable therapeutic strategy against TNBC.

Reciprocal regulation of TLR2-mediated IFN-\u03b2 production by SHP2 and Gsk3\u03b2

Toll-like receptor 2 (TLR2) mediates the innate immune response to bacterial lipopeptides and peptidoglycans by stimulating the production of inflammatory cytokines. However, the mechanisms by which TLR2 signaling regulates type I interferon (IFN)-β production are poorly understood. Here, we identified Src homology 2-containing protein tyrosine phosphatase 2 (SHP2) as a negative regulator of TLR2-induced IFN-β production. Pharmacological inhibition or reduced expression of SHP2 potentiated TLR2 agonist-mediated IFN-β transcription and STAT1 activation, whereas overexpression of SHP2 impaired IFN-β transcription and STAT1 activation. SHP2 physically associated with the glycogen synthase kinase 3β (Gsk3β) in an agonist-dependent manner. Gsk3β positively regulates transcription of IFN-β following TLR2 stimulation by inhibiting the phosphorylation of SHP2. SHP2 inhibited the transcriptional activity of IRF-1 and IRF-8 at the IFN-β promoter. Remarkably, IRF-1 and IRF-8 are recruited to the IFN-β promoter in a SHP2 phosphatase activity-dependent manner. These findings provide insight into the mechanisms by which SHP2 and Gsk3β work together to modulate TLR2-mediated IFN-β production in macrophages.

A Novel Role for RAR\u03b1 Agonists as Apolipoprotein CIII Inhibitors Identified from High Throughput Screening

Elevated triglyceride (TG) levels are well-correlated with the risk for cardiovascular disease (CVD). Apolipoprotein CIII (ApoC-III) is a key regulator of plasma TG levels through regulation of lipolysis and lipid synthesis. To identify novel regulators of TG levels, we carried out a high throughput screen (HTS) using an ApoC-III homogenous time resolved fluorescence (HTRF) assay. We identified several retinoic acid receptor (RAR) agonists that reduced secreted ApoC-III levels in human hepatic cell lines. The RARα specific agonist AM580 inhibited secreted ApoC-III by >80% in Hep3B cells with an EC50 ~2.9 nM. In high-fat diet induced fatty-liver mice, AM580 reduced ApoC-III levels in liver as well as in plasma (~60%). In addition, AM580 treatment effectively reduced body weight, hepatic and plasma TG, and total cholesterol (TC) levels. Mechanistically, AM580 suppresses ApoC-III synthesis by downregulation of HNF4α and upregulation of SHP1 expression. Collectively, these studies suggest that an RARα specific agonist may afford a new strategy for lipid-lowering and CVD risk reduction.

Abstract 4192: Targeting SHP-1/p-Lyn signaling shows therapeutic potential in diffuse large B-cell lymphoma

Abstract Background: Diffuse large B cell lymphoma (DLBCL) is one aggressive form of non-Hodgkin’s lymphoma. Genetic analyses revealed molecular heterogeneity of DLBCL tumors, classifying the cell-of-origin into two distinct molecular subtypes: germinal center B-cell (GCB) and activated B-cell (ABC). ABC-type DLBCL has a worse survival after upfront chemotherapy compared to GCB-type DLBCL, thus ABC-type DLBCL patients have an unmet medical need that warrants additional research efforts and new therapeutic options. In current study we explored the biological role and potential therapeutic implication of a protein phosphatase SHP-1 in DLBCL. Methods: DLBCL cell lines including ABC-like cell lines U2932, Ly-3 and GC-like cell lines DHL-6, Ly-7 and DB were used for in vitro studies. Cell viability was examined by MTT assay. Apoptotic effects were examined by flow cytometry and Western blot. Signal transduction pathways in cells were assessed by Western blot. In vivo therapeutic testing of SHP-1 agonists were performed in nude mice with DLBCL xenografts. Results: We first examined the protein expression of SHP-1 and its downstream p-STAT3 in a panel of DLBCL cell lines, and identified in general SHP-1/p-STAT3 expression was higher in ABC-like cells. Interestingly, the expressions of p-Lyn (Tyr396), p-BTK (Tyr223), key members of B-cell receptor (BCR) signaling pathway, were also higher in ABC-like cells. Knockdown or overexpression of SHP-1 protein expression revealed a reciprocal change of p-Lyn, suggesting SHP-1 negatively regulates phosphorylation of Lyn kinase. Immunoprecipitation experiments confirmed SHP-1 interact with Lyn in DLBCL cells. We previously developed direct SHP-1 agonists, namely SC-43 and SC-60, which could increase SHP-1 activities and induce apoptosis. Here we tested SC-43 and SC-60 in comparison to ibrutinib, a selective Bruton's tyrosine kinase (BTK) inhibitor. The SHP-1 agonists showed in general superior anti-proliferative and apoptotic effects, comparing to ibrutinib. Mechanistically, SHP-1 agonists enhanced SHP-1 activity, decreased BCR signaling p-Lyn and p-BTK, which led to apoptosis. In addition, SHP-1 agonists also down-regulated p-STAT3 as previously reported, which also contributes to anti-cancer effects. In vivo, SC-43 at doses of 10mg/kg/day and 30mg/kg/day orally showed comparable anti-tumor effects with ibrutinib at doses of 12.5 mg/kg/day and 25 mg/kg/day in mice bearing U2932 xenografts, respectively. Western blot confirmed SC-43 downregulation on p-Lyn and p-BTK in vivo. Conclusions: SHP-1 negatively regulates phosphorylation of Lyn, and targeting SHP-1/p-Lyn with direct SHP-1 agonists shows therapeutic potential in DLBCL. Citation Format: Chun-Yu Liu, Man-Hsin Hung, Ming-Hsien Tsai, Pei-Yi Chu, Tzu-Ting Huang, Chun-Teng Huang, Chung-Wai Shiau, Kuen-Feng Chen. Targeting SHP-1/p-Lyn signaling shows therapeutic potential in diffuse large B-cell lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4192. doi:10.1158/1538-7445.AM2017-4192

SHP2 negatively regulates HLA-ABC and PD-L1 expression via STAT1 phosphorylation in prostate cancer cells.

Src homology region 2-containing protein tyrosine phosphatase 2 (SHP2) is a ubiquitous protein tyrosine phosphatase that activates the signal transduction pathways of several growth factors and cytokines. In our study, SHP2 expression was very high in prostate cancer (PCa) cell lines, and the expression of phospho-signal transducer and activator of transcription 1 (p-STAT1) and STAT1 was very low. SHP2 knockdown upregulated the expression of p-STAT1 and downregulated phospho-extracellular signal regulated kinase (p-ERK). SHP2 depletion also increased the expression of human leukocyte antigen (HLA)-ABC and programmed death ligand 1 (PD-L1). When tumor cells were pretreated with Janus kinase 2 (JAK2) inhibitor, SHP2 depletion failed to induce HLA-ABC and PD-L1 expression. Furthermore, treating tumor cells with the mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) inhibitor PD0325901 did not upregulate HLA-ABC and PD-L1. SHP2 depletion was associated with increased T-cell activation (CD25 MFI of CD8+) by coculture of allogeneic healthy donor peripheral blood monocytes (PBMC) with SHP2 siRNA pretreated PCa cell lines. These results show that SHP2 targeting upregulates HLA-ABC and PD-L1 expression via STAT1 phosphorylation in PCa cells and SHP2 depletion could increase T-cell activation.

A pumpkin polysaccharide induces apoptosis by inhibiting the JAK2/STAT3 pathway in human hepatoma HepG2 cells

The purpose of this study is to investigate the effect of a purified polysaccharide (PPPF) from pumpkin fruit on the Janus activated kinase (JAK)/signal transducer and activator of transcription (STAT) signaling during apoptotic process. The results showed that PPPF or STAT3 siRNA inhibits the cell growth of HepG2 cells via induction of apoptosis. Moreover, PPPF is able to suppress both constitutive and IL-6-induced phosphorylation of STAT3 (on Tyr705) and subsequent nuclear translocation in cancer cells. Such inhibition is found to be achieved through down-regulation of constitutive phosphorylation of JAK2, but not JAk1, c-Src, ERK1/2, and Akt, which means STAT3 tyrosine phosphorylation in HepG2 cells following PPPF treatment is associated with a reduction in JAK2 activity. In addition, the protein expression of SHP-1 was increased in cells in response to PPPF treatment, whereas SHP-2, SOCS-1 and SCOS-3 protein expression remain unchanged. In vivo animal experiment also indicated that PPPF had a potent inhibitory effect on tumor growth in mice bearing HepG2 xenograft tumors. Thus we can conclude that PPPF directly induces apoptotic cell death of HepG2 cells via down-regulation of the JAK2/STAT3 signal transduction pathways, which may facilitates the development of a therapeutic strategy for treating HCC.

Role and mechanism of decitabine combined with tyrosine kinase inhibitors in advanced chronic myeloid leukemia cells.

Patients with advanced chronic myeloid leukemia (CML) have a poor prognosis, with the use of tyrosine kinase inhibitors (TKIs) to treat CML demonstrating poor results. The results of the present study revealed that, following Cell Counting Kit-8 analysis, treatment of K562 cells with decitabine (DAC) combined with TKIs exhibits synergic effects. Co-immunoprecipitation indicated that tyrosine-protein phosphatase non-receptor type 6 (SHP-1) and BCR-ABL fusion protein (BCR-ABL) (p210) form a complex in the K562 cell line, and in the primary cells derived from patients with CML. These results suggested that SHP-1 serves a role in regulating the tyrosine kinase activity of BCR-ABL (p210). In addition, SHP-1 expression increased, while BCR-ABL expression decreased in the group treated with DAC and TKIs combined group compared with the TKI monotherapy group. Treatment with imatinib (IM) demonstrated no effect on SHP-1 methylation in the K562 cell line; however, the methylation of SHP-1 was not determined in the combined IM and DAC therapy group. Treatment with DAC demonstrated the ability to activate the expression of silenced SHP-1 through demethylation, thus decreasing BCR-ABL tyrosine kinase activity, resulting in an improved therapeutic effect on CML.

Omega-3 free fatty acids attenuate insulin-promoted breast cancer cell proliferation

High insulin levels in obese people are considered as a risk factor to induce breast carcinogenesis. And consumption of fish oils which mainly contain omega-3 fatty acids is associated with a reduced risk of breast cancer. However, whether omega-3 free fatty acids (FFAs) modulate insulin signaling pathway to prevent breast cancer is poorly understood. The current study tested the hypothesis that omega-3 FFAs attenuate insulin-induced breast cancer cell proliferation and regulate insulin signaling pathway. We show here that omega-3 FFAs attenuate MCF-7 cell proliferation and Akt and Erk1/2 phosphorylation levels stimulated by insulin. Knockdown Shp2 by siRNA resulted in significantly elevated omega-3 FFAs–activated Akt phosphorylation but failed to change insulin-stimulated Akt and Erk1/2 phosphorylation. And viable cell number was not affected by either downregulation of Shp2 expression or Erk1/2 inhibitor U0126 treatment. These observations indicated that omega-3 FFAs attenuate insulin-promoted breast cancer cell proliferation and insulin-activated Akt phosphorylation.

Crocin Suppresses Constitutively Active STAT3 Through Induction of Protein Tyrosine Phosphatase SHP‐1

The aim of the present study is to investigate the effect of a natural compound crocin, one of the active components of saffron, on human multiple myeloma cells. Crocin effectively suppressed constitutive STAT3 activation, translocation of STAT3 to the nucleus, and its target gene expression. The suppression of STAT3 was mediated through the inhibition of activation of protein tyrosine kinases JAK1, JAK2, and c-Src. We found that crocin induced the expression of SHP-1, a tyrosine protein phosphatase, and pervanadate treatment reversed the crocin-induced downregulation of STAT3, suggesting the involvement of a protein tyrosine phosphatase. Moreover, suppression of SHP-1 by its inhibitor overturned the effect of crocin on induction of SHP-1 and the inhibition of STAT3 activation. Finally, crocin downregulated the expression of STAT3-mediated gene products including anti-apoptotic (Bcl-2), pro-apoptotic (BAX), invasive (CXCR4), angiogenic (VEGF), and cell cycle regulator (cyclin D1), which are correlated with suppression of proliferation, the accumulation of cells in sub-G1 phase of cell cycle, and induction of apoptosis. Overall, our results suggested that crocin is a novel inhibitor of STAT3 activation pathway and thus may have potential in prevention and treatment of human multiple myeloma. J. Cell. Biochem. 118: 3290-3298, 2017. © 2017 Wiley Periodicals, Inc.

Shp1 positively regulates EGFR signaling by controlling EGFR protein expression in mammary epithelial cells

SH2-domain containing protein tyrosine phosphatase 1 (Shp1/PTPN6) is mainly expressed in hematopoietic cells and acts a negative signaling regulator. Although Shp1 is also expressed in epithelial cells, the function of shp1 in normal epithelial is still less well understood, especially in regulating the growth of epithelial cells. In this study, different shRNAs and siRNAs against Shp1 were used to knockdown Shp1 expression in MCF10A, an immortalized mammary epithelial cell line. Shp1 knockdown resulted in inhibited cell growth in part due to lower percentage of MCF10A cells entering into S phase and reduced cyclin D1 expression. Accordingly, EGF-induced tyrosyl phosphorylation of EGFR and Stat5 was significantly inhibited in cells with Shp1 knockdown compared with control whereas EGF-induced Akt and Erk phosphorylation was not affected by Shp1 knockdown. Further analysis revealed that Shp1 knockdown lead to decreased EGFR protein expression without affecting EGFR mRNA expression or increasing EGFR protein degradation. Our data indicate that Shp1 functions as a positive regulator and acts in a novel mechanism through promoting EGFR protein expression in mammary epithelial cells.

Inhibition of Src homology 2 domain containing protein tyrosine phosphatase as the possible mechanism of metformin-assisted amelioration of obesity induced insulin resistance in high fat diet fed C57BL/6J mice

SHP-1 (Src homology 2 domain containing protein tyrosine phosphatase) is a known negative regulator of insulin signaling and inflammation. To date, the molecular mechanism of metformin in modulating SHP-1 expression has remained elusive. In the present study, we have investigated the role of SHP-1 in relation to anti-hyperglycemic and anti-inflammatory actions of metformin in an obese phenotype mouse model. We observed that metformin treatment significantly reduced SHP-1 activity in obese mice, leading to improved insulin sensitivity. Additionally, metformin down regulated inflammatory markers like TLR2, TLR4, CD80, CD86, NF-κB, STAT1 and suppressed adipose tissue inflammation by efficiently polarizing adipose tissue macrophages toward anti-inflammatory state by way of indirect inhibition of SHP-1 mRNA and protein expressions. Our study suggests that metformin exerts its insulin sensitizing effects via inhibition of SHP-1 activity and expression.

Abnormal repression of SHP-1, SHP-2 and SOCS-1 transcription sustains the activation of the JAK/STAT3 pathway and the progression of the disease in multiple myeloma.

Sustained activation of JAK/STAT3 signaling pathway is classically described in Multiple Myeloma (MM). One explanation could be the silencing of the JAK/STAT suppressor genes, through the hypermethylation of SHP-1 and SOCS-1, previously demonstrated in MM cell lines or in whole bone marrow aspirates. The link between such suppressor gene silencing and the degree of bone marrow invasion or the treatment response has not been evaluated in depth. Using real-time RT-PCR, we studied the expression profile of three JAK/STAT suppressor genes: SHP-1, SHP-2 and SOCS-1 in plasma cells freshly isolated from the bone marrows of MM patients and healthy controls. Our data demonstrated an abnormal repression of such genes in malignant plasma cells and revealed a significant correlation between such defects and the sustained activation of the JAK/STAT3 pathway during MM. The repressed expression of SHP-1 and SHP-2 correlated significantly with a high initial degree of bone marrow infiltration but was, unexpectedly, associated with a better response to the induction therapy. Collectively, our data provide new evidences that substantiate the contribution of JAK/STAT suppressor genes in the pathogenesis of MM. They also highlight the possibility that the decreased gene expression of SHP-1 and SHP-2 could be of interest as a new predictive factor of a favorable treatment response, and suggest new potential mechanisms of action of the therapeutic molecules. Whether such defect helps the progression of the disease from monoclonal gammopathy of unknown significance to MM remains, however, to be determined.

Protein tyrosine phosphatase Shp2 deficiency in podocytes attenuates lipopolysaccharide-induced proteinuria

Podocytes are specialized epithelial cells that play a significant role in maintaining the integrity of the glomerular filtration barrier and preventing urinary protein leakage. We investigated the contribution of protein tyrosine phosphatase Shp2 to lipopolysaccharide (LPS)-induced renal injury. We report increased Shp2 expression in murine kidneys and cultured podocytes following an LPS challenge. To determine the role of podocyte Shp2 in vivo, we generated podocyte-specific Shp2 knockout (pod-Shp2 KO) mice. Following administration of LPS, pod-Shp2 KO mice exhibited lower proteinuria and blood urea nitrogen concentrations than controls indicative of preserved filter integrity. In addition, renal mRNA and serum concentrations of inflammatory cytokines IL-1β, TNFα, INFγ and IL-12 p70 were significantly decreased in LPS-treated knockout mice compared with controls. Moreover, the protective effects of podocyte Shp2 deficiency were associated with decreased LPS-induced NF-κB and MAPK activation, nephrin phosphorylation and attenuated endoplasmic reticulum stress. These effects were recapitulated in differentiated E11 murine podocytes with lentiviral-mediated Shp2 knockdown. Furthermore, Shp2 deficient podocytes displayed reduced LPS-induced migration in a wound healing assay. These findings identify Shp2 in podocytes as a significant contributor to the signaling events following LPS challenge and suggest that inhibition of Shp2 in podocytes may present a potential therapeutic target for podocytopathies.

DNA methyltransferase 1 mediated aberrant methylation and silencing of SHP-1 gene in chronic myelogenous leukemia cells

IntroductionExtensive studies on SHP-1 protein and SHP-1 mRNA revealed that the diminishment or abolishment of the expression of SHP-1 in leukemias/lymphomas was due to aberrant promoter methylation. Thus far, the mechanism of epigenetic silencing of the SHP-1 tyrosine phosphatase gene that occurs in chronic myelogenous leukemia cells remains poorly understood. MethodsThe expressions of the target molecules were determined by quantitative real time PCR and western blot, respectively. Bisulfite sequencing PCR was used to detect methylation status of DNA CpG. The lentiviral vectors were applied to modify gene expression. ResultsIn the present study, we found that the promoter 2 of SHP-1 gene is located between positions from −577bp to +300bp, and 22 CpG sites contained in positions −353bp∼+182bp are aberrantly methylated in K562 cells. In vitro, we demonstrated that DNMT1 silencing induced demethylation of the 22 CpG sites located in the SHP-1 promoter and re-expression of SHP-1 gene in K562 cells. Moreover, we proved that the expression levels of DNMT1 and SHP-1 mRNA and protein were negatively correlated in K562 cells and BM aspirates mononuclear cells from CML patients. ConclusionCollectively, these results indicate that DNMT1 mediates aberrant methylation and silencing of SHP-1 gene in chronic myelogenous leukemia cells, and provide a novel therapeutic target for CML.

SHP2 is induced by the HBx-NF-κB pathway and contributes to fibrosis during human early hepatocellular carcinoma development.

The non-receptor tyrosine phosphatase SHP2 has scaffolding functions in signal transduction cascades downstream of growth receptors. A recent study suggested that SHP2 acts as a tumor suppressor during hepatocellular carcinoma (HCC) development. Herein we examined whether SHP2 links the HBx–NF-κB pathway to EGFR signaling during HCC development. The overexpression of HBx or NF-κB led to increased SHP2 expression via NF-κB binding to the Shp2 promoter. EGF treatment induced ERK activation as well as the rapid assembly of SHP2, EGFR, and Gab1. Upon LPS stimulation, NF-κB–SHP2–ERK activation and phosphorylated STAT3 levels exhibited a negative correlation in vitro. By contrast, in patients with HBV-associated HCC, NF-κB–SHP2–ERK and IL-6–JAK–STAT3 pathway activity levels were concomitantly higher in adjacent non-neoplastic tissues than in HCC tissues. The immunohistochemical analysis of 162 tissues of patients with HCC revealed that SHP2 levels were significantly higher in non-neoplastic background tissues than in corresponding HCC tissues and considerably increased in background liver tissues with advanced fibrosis (P < 0.001). SHP2 expression increased gradually from normal liver to chronic hepatitis, cirrhosis, and background liver with a dysplastic nodule, but was decreased or lost in dysplastic nodules and HCC. This is the first report to describe the existence of the HBx–NF-κB–SHP2 pathway, linking HBV infection to the EGFR–RAS–RAF–MAPK pathway in the liver. SHP2 depletion from the negative crosstalk between NF-κB and STAT3 accelerates HCC development.

Expression of SHP-1 and SOCS6 in patients with acute leukemia and their clinical implication.

BackgroundTo investigate the expression and clinical relevance of Src homology region 2 domain-containing phosphatase-1 (SHP-1) and suppressor of cytokine signaling 6 (SOCS6) in acute leukemia (AL).Patients and methodsThe enrolled AL patients were divided into three groups (newly diagnosed, relapsed, and complete remission [CR]). Healthy donors were also included as a control group in this study. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) was performed to measure messenger RNA (mRNA) expression of SHP-1 and SOCS6. Statistical analysis was conducted to analyze the correlation between mRNA levels of SHP-1 and SOCS6 with patient outcomes.ResultsmRNA expression of SHP-1 was significantly lower in AL patients than that in healthy donors. The newly diagnosed or relapsed AL patients had lower mRNA levels of SHP-1 than the patients in CR. In contrast, SOCS6 mRNA expression was significantly higher in newly diagnosed or relapsed patients than that in patients in CR as well as healthy donors. However, mRNA levels of both SHP-1 and SOCS6 were positively correlated with the patient remission. The chemotherapy-induced remission rate was higher in patients with detectable SHP-1 or SOCS6 expression than in patients with undetectable SHP-1 or SOCS6 expression. Furthermore, the AL patients with detectable SHP-1 mRNA expression had lower incidence rate of invasive fungal infection.ConclusionThe results suggest that expression patterns of SHP-1 and SOCS6 differ in AL patients. Despite the difference, expression of SHP-1 and SOCS6 is associated with favorable outcomes, suggesting an anticancer property of these two genes in AL.

Ginkgolic Acid C 17:1, Derived from Ginkgo biloba Leaves, Suppresses Constitutive and Inducible STAT3 Activation through Induction of PTEN and SHP-1 Tyrosine Phosphatase.

Ginkgolic acid C 17:1 (GAC 17:1) extracted from Ginkgo biloba leaves, has been previously reported to exhibit diverse antitumor effect(s) through modulation of several molecular targets in tumor cells, however the detailed mechanism(s) of its actions still remains to be elucidated. Signal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor that regulates various critical functions involved in progression of diverse hematological malignancies, including multiple myeloma, therefore attenuating STAT3 activation may have a potential in cancer therapy. We determined the anti-tumor mechanism of GAC 17:1 with respect to its effect on STAT3 signaling pathway in multiple myeloma cell lines. We found that GAC 17:1 can inhibit constitutive activation of STAT3 through the abrogation of upstream JAK2, Src but not of JAK1 kinases in U266 cells and also found that GAC can suppress IL-6-induced STAT3 phosphorylation in MM.1S cells. Treatment of protein tyrosine phosphatase (PTP) inhibitor blocked suppression of STAT3 phosphorylation by GAC 17:1, thereby indicating a critical role for a PTP. We also demonstrate that GAC 17:1 can induce the substantial expression of PTEN and SHP-1 at both protein and mRNA level. Further, deletion of PTEN and SHP-1 genes by siRNA can repress the induction of PTEN and SHP-1, as well as abolished the inhibitory effect of drug on STAT3 phosphorylation. GAC 17:1 down-regulated the expression of STAT3 regulated gene products and induced apoptosis of tumor cells. Overall, GAC 17:1 was found to abrogate STAT3 signaling pathway and thus exert its anticancer effects against multiple myeloma cells.