Integrin αvß3 Upregulation in Response to Nutrient Stress Promotes Lung Cancer Cell Metabolic Plasticity.

Cancer stem/tumor-initiating cells display stress tolerance and metabolic flexibility to survive in a harsh environment with limited nutrient and oxygen availability. The molecular mechanisms underlying this phenomenon could provide targets to prevent metabolic adaptation and halt cancer progression. Here, we showed in cultured cells and live human surgical biopsies of non-small cell lung cancer that nutrient stress drives the expression of the epithelial cancer stem cell marker integrin αvß3 via upregulation of the ß3 subunit, resulting in a metabolic reprogramming cascade that allows tumor cells to thrive despite a nutrient-limiting environment. Although nutrient deprivation is known to promote acute, yet transient, activation of the stress sensor AMP-activated protein kinase (AMPK), stress-induced αvß3 expression via Src activation unexpectedly led to secondary and sustained AMPK activation. This resulted in the nuclear localization of peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC1α) and upregulation of glutamine metabolism, the tricarboxylic acid cycle, and oxidative phosphorylation. Pharmacological or genetic targeting of this axis prevented lung cancer cells from evading the effects of nutrient stress, thereby blocking tumor initiation in mice following orthotopic implantation of lung cancer cells. These findings reveal a molecular pathway driven by nutrient stress that results in cancer stem cell reprogramming to promote metabolic flexibility and tumor initiation. SIGNIFICANCE: Upregulation of integrin αvß3, a cancer stem cell marker, in response to nutrient stress activates sustained AMPK/PGC1α signaling that induces metabolic reprogramming in lung cancer cells to support their survival. See related commentary by Rainero, p. 1543.

Mechanical Rippling for Diverse Ferroelectric Topologies in Otherwise Nonferroelectric SrTiO_{3} Nanofilms.

Polar topological structures such as skyrmions and merons have become an emerging research field due to their rich functionalities and promising applications in information storage. Up to now, the obtained polar topological structures are restricted to a few limited ferroelectrics with complex heterostructures, limiting their large-scale practical applications. Here, we circumvent this limitation by utilizing a nanoscale ripple-generated flexoelectric field as a universal means to create rich polar topological configurations in nonpolar nanofilms in a controllable fashion. Our extensive phase-field simulations show that a rippled SrTiO_{3} nanofilm with a single bulge activates polarizations that are stabilized in meron configurations, which further undergo topological transitions to Néel-type and Bloch-type skyrmions upon varying the geometries. The formation of these topologies originates from the curvature-dependent flexoelectric field, which extends beyond the common mechanism of geometric confinement that requires harsh energy conditions and strict temperature ranges. We further demonstrate that the rippled nanofilm with three-dimensional ripple patterns can accommodate other unreported modulated phases of ferroelectric topologies, which provide ferroelectric analogs to the complex spin topologies in magnets. The present study not only unveils the intriguing nanoscale electromechanical properties but also opens exciting opportunities to design various functional topological phenomena in flexible materials.

Upregulation of fibronectin and its integrin receptors - an adaptation to isolation stress that facilitates tumor initiation.

Tumor initiation at either primary or metastatic sites is an inefficient process in which tumor cells must fulfill a series of conditions. One critical condition involves the ability of individual tumor-initiating cells to overcome 'isolation stress', enabling them to survive within harsh isolating microenvironments that can feature nutrient stress, hypoxia, oxidative stress and the absence of a proper extracellular matrix (ECM). In response to isolation stress, tumor cells can exploit various adaptive strategies to develop stress tolerance and gain stemness features. In this Opinion, we discuss how strategies such as the induction of certain cell surface receptors and deposition of ECM proteins enable tumor cells to endure isolation stress, thereby gaining tumor-initiating potential. As examples, we highlight recent findings from our group demonstrating how exposure of tumor cells to isolation stress upregulates the G-protein-coupled receptor lysophosphatidic acid receptor 4 (LPAR4), its downstream target fibronectin and two fibronectin-binding integrins, α5ß1 and αvß3. These responses create a fibronectin-rich niche for tumor cells, ultimately driving stress tolerance, cancer stemness and tumor initiation. We suggest that approaches to prevent cancer cells from adapting to stress by suppressing LPAR4 induction, blocking its downstream signaling or disrupting fibronectin-integrin interactions hold promise as potential strategies for cancer treatment.

Tumor-initiating cells establish a niche to overcome isolation stress.

While the tumor microenvironment is a critical contributor to cancer progression, early steps of tumor initiation and metastasis also rely on the ability of individual tumor cells to survive and thrive at locations where tumor stroma or immune infiltration has yet to be established. In this opinion article, we use the term 'isolation stress' to broadly describe the challenges that individual tumor cells must overcome during the initiation and expansion of the primary tumor beyond permissive boundaries and metastatic spread into distant sites, including a lack of cell-cell contact, adhesion to protumor extracellular matrix proteins, and access to nutrients, oxygen, and soluble factors that support growth. In particular, we highlight the ability of solitary tumor cells to autonomously generate a specialized fibronectin-enriched extracellular matrix to create their own pericellular niche that supports tumor initiation. Cancer cells that can creatively evade the effects of isolation stress not only become more broadly stress tolerant, they also tend to show enhanced stemness, drug resistance, tumor initiation, and metastasis.

Pancreatic cancer cells upregulate LPAR4 in response to isolation stress to promote an ECM-enriched niche and support tumour initiation.

Defining drivers of tumour initiation can provide opportunities to control cancer progression. Here we report that lysophosphatidic acid receptor 4 (LPAR4) becomes transiently upregulated on pancreatic cancer cells exposed to environmental stress or chemotherapy where it promotes stress tolerance, drug resistance, self-renewal and tumour initiation. Pancreatic cancer cells gain LPAR4 expression in response to stress by downregulating a tumour suppressor, miR-139-5p. Even in the absence of exogenous lysophosphatidic acid, LPAR4-expressing tumour cells display an enrichment of extracellular matrix genes that are established drivers of cancer stemness. Mechanistically, upregulation of fibronectin via an LPAR4/AKT/CREB axis is indispensable for LPAR4-induced tumour initiation and stress tolerance. Moreover, ligation of this fibronectin-containing matrix via integrins α5ß1 or αVß3 can transfer stress tolerance to LPAR4-negative cells. Therefore, stress- or drug-induced LPAR4 enhances cell-autonomous production of a fibronectin-rich extracellular matrix, allowing cells to survive 'isolation stress' and compensate for the absence of stromal-derived factors by creating their own tumour-initiating niche.

Metabolic Reprograming via Deletion of CISH in Human iPSC-Derived NK Cells Promotes In Vivo Persistence and Enhances Anti-tumor Activity.

Cytokine-inducible SH2-containing protein (CIS; encoded by the gene CISH) is a key negative regulator of interleukin-15 (IL-15) signaling in natural killer (NK) cells. Here, we develop human CISH-knockout (CISH-/-) NK cells using an induced pluripotent stem cell-derived NK cell (iPSC-NK cell) platform. CISH-/- iPSC-NK cells demonstrate increased IL-15-mediated JAK-STAT signaling activity. Consequently, CISH-/- iPSC-NK cells exhibit improved expansion and increased cytotoxic activity against multiple tumor cell lines when maintained at low cytokine concentrations. CISH-/- iPSC-NK cells display significantly increased in vivo persistence and inhibition of tumor progression in a leukemia xenograft model. Mechanistically, CISH-/- iPSC-NK cells display improved metabolic fitness characterized by increased basal glycolysis, glycolytic capacity, maximal mitochondrial respiration, ATP-linked respiration, and spare respiration capacity mediated by mammalian target of rapamycin (mTOR) signaling that directly contributes to enhanced NK cell function. Together, these studies demonstrate that CIS plays a key role to regulate human NK cell metabolic activity and thereby modulate anti-tumor activity.

Decoration of Anti-CD38 on Nanoparticles Carrying a STAT3 Inhibitor Can Improve the Therapeutic Efficacy Against Myeloma.

STAT3 is an oncoprotein which has been shown to contribute to drug resistance in multiple myeloma (MM). Nonetheless, the clinical utility of STAT3 inhibitors in treating MM has been limited, partly related to some of their pharmacologic properties. To overcome these challenges, our group had previously packaged STAT3 inhibitors using a novel formulation of nanoparticles (NP) and found encouraging results. In this study, we aimed to further improve the pharmacologic properties of these NP by decorating them with monoclonal anti-CD38 antibodies. NP loaded with S3I-1757 (a STAT3 inhibitor), labeled as S3I-NP, were generated. S3I-NP decorated with anti-CD38 (labeled as CD38-S3I-NP) were found to have a similar nanoparticular size, drug encapsulation, and loading as S3I-NP. The release of S3I-1757 at 24 h was also similar between the two formulations. Using Cy5.5 labeling of the NP, we found that the decoration of anti-CD38 on these NP significantly increased the cellular uptake by two MM cell lines (p < 0.001). Accordingly, CD38-S3I-NP showed a significantly lower inhibitory concentration at 50% (IC50) compared to S3I-NP in two IL6-stimulated MM cell lines (p < 0.001). In a xenograft mouse model, CD38-S3I-NP significantly reduced the tumor size by 4-fold compared to S3I-NP on day 12 after drug administration (p = 0.006). The efficacy of CD38-S3I-NP in suppressing STAT3 phosphorylation in the xenografts was confirmed by using immunocytochemistry and Western blot analysis. In conclusion, our study suggests that the decoration of anti-CD38 on NP loaded with STAT3 inhibitors can further improve their therapeutic effects against MM.

Oxidative stress enhances tumorigenicity and stem-like features via the activation of the Wnt/ß-catenin/MYC/Sox2 axis in ALK-positive anaplastic large-cell lymphoma.

BACKGROUND: The phenomenon that malignant cells can acquire stemness under specific stimuli, encompassed under the concept of cancer cell plasticity, has been well-described in epithelial malignancies. To our knowledge, cancer cell plasticity has not yet been described in hematopoietic cancers. To illustrate and study cancer cell plasticity in hematopoietic cancers, we employed an in-vitro experimental model of ALK-positive anaplastic large-cell lymphoma (ALK+ALCL) that is based on the phenotypic and functional dichotomy of these cells, with cells responsive to a Sox2 reporter (i.e. RR cells) being significantly more stem-like than those unresponsive to the reporter (i.e. RU cells). METHODS: H2O2 was employed to trigger oxidative stress. GFP expression and luciferase activity, readouts of the Sox2 reporter activity, were quantified by using flow cytometry and luciferase activity assay, respectively. Doxorubicin-resistance and clonogenicity were assessed by using the MTS, methylcellulose colony formation and limiting dilution assays. Western blotting and quantitative PCR were used to assess the expression of various members of the Wnt/ß-catenin pathway. Pull-down studies using a Sox2 binding consensus sequence were used to assess Sox2-DNA binding. Quercetin and 10074-G5 were used to inhibit ß-catenin and MYC, respectively. siRNA was used to downregulate Sox2. RESULTS: Under H2O2-induced oxidative stress, a substantial fraction of RU cells was found to convert to RR cells, as evidenced by their acquisition of GFP expression and luciferase activity. Compared to the native RU cells, converted RR cells had significantly higher levels of doxorubicin-resistance, clonogenicity and sphere formation. Converted RR cells were characterized by an upregulation of the Wnt/ß-catenin/MYC/Sox2 signaling axis, previously found to be the key regulator of the RU/RR dichotomy in ALK+ALCL. Furthermore, Sox2 was found to bind to DNA efficiently in converted RR cells but not RU cells, and this finding correlated with significant elevations of several Sox2 downstream targets such as WNT2B and BCL9. Lastly, inhibition of ß-catenin, MYC or Sox2 in RU cells significantly abrogated the H2O2-induced RU/RR conversion. CONCLUSIONS: We have demonstrated that cancer cell plasticity exists in ALK+ALCL, a type of hematopoietic cancer. In this cancer type, the Wnt/ß-catenin/MYC/Sox2 axis is an important regulator of cancer cell plasticity.

Phosphorylation of Sox2 at Threonine 116 is a Potential Marker to Identify a Subset of Breast Cancer Cells with High Tumorigenecity and Stem-Like Features.

We have previously identified a novel phenotypic dichotomy in breast cancer (BC) based on the response to a SRR2 (Sox2 regulatory region 2) reporter, with reporter responsive (RR) cells being more tumorigenic/stem-like than reporter unresponsive (RU) cells. Since the expression level of Sox2 is comparable between the two cell subsets, we hypothesized that post-translational modifications of Sox2 contribute to their differential reporter response and phenotypic differences. By liquid chromatography-mass spectrometry, we found Sox2 to be phosphorylated in RR but not RU cells. Threonine 116 is an important phosphorylation site, since transfection of the T116A mutant into RR cells significantly decreased the SRR2 reporter luciferase activity and the RR-associated phenotype. Oxidative stress-induced conversion of RU into RR cells was accompanied by Sox2 phosphorylation at T116 and increased Sox2-DNA binding. In a cohort of BC, we found significant correlations between the proportion of tumor cells immuno-reactive with anti-phosphorylated Sox2T116 and a high tumor grade (p = 0.006), vascular invasion (p = 0.001) and estrogen receptor expression (p = 0.032). In conclusion, our data suggests that phosphorylation of Sox2T116 contributes to the tumorigenic/stem-like features in RR cells. Detection of phospho-Sox2T116 may be useful in identifying a small subset of tumor cells carrying stem-like/tumorigenic features in BC.

High expression of ß-catenin contributes to the crizotinib resistant phenotype in the stem-like cell population in neuroblastoma.

ALK has been identified as a novel therapeutic target in neuroblastoma (NB), but resistance to ALK inhibitors (such as crizotinib) is well recognized. We recently published that the crizotinib sensitivity in NB cells strongly correlates with the crizotinib-ALK binding, and ß-catenin effectively hinders this interaction and confers crizotinib resistance. Here, we asked if these observations hold true for the stem-like cells in NB cells, which were purified based on their responsiveness to a Sox2 reporter. Compared to bulk, reporter unresponsive (RU) cells, reporter responsive (RR) cells had significantly higher neurosphere formation ability, expression of CD133/nestin and chemo-resistance. Using the cellular thermal shift assay, we found that RR cells exhibited significantly weaker crizotinib-ALK binding and higher crizotinib resistance than RU cells. The suboptimal crizotinib-ALK binding in RR cells can be attributed to their high ß-catenin expression, since siRNA knockdown of ß-catenin restored the crizotinib-ALK binding and lowered the crizotinib resistance to the level of RU cells. Enforced expression of ß-catenin in RU cells resulted in the opposite effects. To conclude, high expression of ß-catenin in the stem-like NB cells contributes to their crizotinib resistance. Combining ß-catenin inhibitors and ALK inhibitors may be useful in treating NB patients.

[Research development and thinking of clinical study of Polygonum multiflorum].

In order to offer the accordance and research ideas for further research of effect, indication and ensure the safety and effectiveness of clinical medication, the current states of clinical application and adverse reaction of Polygonum multiflorum were summarized. The related literatures in recent years were collated, analyzed and summed up. P. multiflorum is widely used and has variety of adverse reactions. At present, it's used to treat hyperlipidemia, white hair, hair loss, hypertension, vitiligo, etc. Adverse reactions are liver damage, allergic reactions, mental symptoms, chronic enteritis, etc. The effect, indication and adverse reaction research of P. multiflorum is still insufficient, further study should be based on clinical practice, in the aspects of the authenticity of the drug, the disease syndrome, the phase of adverse reactions, individual heterogeneity and compatibility.

High Myc expression and transcription activity underlies intra-tumoral heterogeneity in triple-negative breast cancer.

We have previously identified a novel intra-tumoral dichotomy in triple-negative breast cancer (TNBC) based on the differential responsiveness to a reporter containing the Sox2 regulatory region-2 (SRR2), with reporter responsive (RR) cells being more stem-like than reporter unresponsive (RU) cells. Using bioinformatics, we profiled the protein-DNA binding motifs of SRR2 and identified Myc as one of the potential transcription factors driving SRR2 activity. In support of its role, Myc was found to be highly expressed in RR cells as compared to RU cells. Enforced expression of MYC in RU cells resulted in a significant increase in SRR2 activity, Myc-DNA binding, proportion of cellsexpressing CD44+/CD24-, chemoresistance and mammosphere formation. Knockdown of Myc using siRNA in RR cells led to the opposite effects. We also found evidence that the relatively high ERK activation in RR cells contributes to their high expression of Myc and stem-like features. Using confocal microscopy and patient samples, we found a co-localization between Myc and CD44 in the same cell population. Lastly, a high proportion of Myc-positive cells in tumors significantly correlated with a short patient survival. In conclusion, inhibition of the MAPK/ERK/Myc axis may be an effective approach in eliminating stem-like cells in TNBC.

A positive feedback loop involving the Wnt/ß-catenin/MYC/Sox2 axis defines a highly tumorigenic cell subpopulation in ALK-positive anaplastic large cell lymphoma.

BACKGROUND: We have previously described the existence of two phenotypically distinct cell subsets in ALK-positive anaplastic large cell lymphoma (ALK + ALCL) based on their differential responsiveness to a Sox2 reporter (SRR2), with reporter-responsive (RR) cells being more tumorigenic and chemoresistant than reporter-unresponsive (RU) cells. However, the regulator(s) of RU/RR dichotomy are not identified. In this study, we aim to delineate the key regulator(s) of RU/RR dichotomy. METHODS: JASPER motif match analysis was used to identify the putative factors binding to SRR2 sequence. SRR2 probe pull-down assay and quantitate real-time PCR were performed to analyze the regulation of Sox2 transcriptional activity by MYC. Methylcellulose colony formation assay, chemoresistance to doxorubicin and mouse xenograft study were performed to investigate the biological functions of MYC. PCR array and western blotting were executed to study related signaling pathways that regulate MYC expression. Immunofluorescence and immunohistochemistry assay were initiated to evaluate the expression of MYC and its correlation with its regulator by chi-square test analysis in human primary tumor cells. RESULTS: We identified MYC as a potential regulator of RU/RR dichotomy. In support of its role, MYC was highly expressed in RR cells compared to RU cells, and inhibition of MYC substantially decreased the Sox2/SRR2 binding, Sox2 transcriptional activity, chemoresistance, and methylcellulose colony formation. In contrast, enforced expression of MYC in RU cells conferred the RR phenotype. The Wnt/ß-catenin pathway, a positive regulator of MYC, was highly active in RR but not RU cells. While inhibition of this pathway in RR cells substantially decreased MYC expression and SRR2 reporter activity, experimental activation of this pathway led to the opposite effects in RU cells. Collectively, our results support a model in which a positive feedback loop involving Wnt/ß-catenin/MYC and Sox2 contributes to the RR phenotype. In a mouse xenograft model, RU cells stably transfected with MYC showed upregulation of the Wnt/ß-catenin/MYC/Sox2 axis and increased tumorigenecity. Correlating with these findings, there was a significant correlation between the expression of active ß-catenin and MYC in ALK + ALCL primary tumor cells. CONCLUSIONS: A positive feedback loop involving the Wnt/ß-catenin/MYC/Sox2 axis defines a highly tumorigenic cell subset in ALK + ALCL.

The use of cellular thermal shift assay (CETSA) to study Crizotinib resistance in ALK-expressing human cancers.

Various forms of oncogenic ALK proteins have been identified in various types of human cancers. While Crizotinib, an ALK inhibitor, has been found to be therapeutically useful against a subset of ALK(+) tumours, clinical resistance to this drug has been well recognized and the mechanism of this phenomenon is incompletely understood. Using the cellular thermal shift assay (CETSA), we measured the Crizotinib-ALK binding in a panel of ALK(+) cell lines, and correlated the findings with the ALK structure and its interactions with specific binding proteins. The Crizotinib IC50 significantly correlated with Crizotinib-ALK binding. The suboptimal Crizotinib-ALK binding in Crizotinib-resistant cells is not due to the cell-specific environment, since transfection of NPM-ALK into these cells revealed substantial Crizotinib-NPM-ALK binding. Interestingly, we found that the resistant cells expressed higher protein level of ß-catenin and siRNA knockdown restored Crizotinib-ALK binding (correlated with a significant lowering of IC50). Computational analysis of the crystal structures suggests that ß-catenin exerts steric hindrance to the Crizotinib-ALK binding. In conclusion, the Crizotinib-ALK binding measurable by CETSA is useful in predicting Crizotinib sensitivity, and Crizotinib-ALK binding is in turn dictated by the structure of ALK and some of its binding partners.

miR-200b induces cell cycle arrest and represses cell growth in esophageal squamous cell carcinoma.

miR-200b is a pleiotropically acting microRNA in cancer progression, representing an attractive therapeutic target. We previously identified miR-200b as an invasiveness repressor in esophageal squamous cell carcinoma (ESCC), whereas further understanding is warranted to establish it as a therapeutic target. Here, we show that miR-200b mitigates ESCC cell growth by inducing G2-phase cell cycle arrest and apoptosis. The expression/activation of multiple key cell cycle regulators such as CDK1, CDK2, CDK4 and Cyclin B, and the Wnt/ß-Catenin signaling are modulated by miR-200b. We identified CDK2 and PAF (PCNA-associated factor), two important tumor-promoting factors, as direct miR-200b targets in ESCC. Correlating with the frequent loss of miR-200b in ESCC, both CDK2 and PAF levels are significantly increased in ESCC tumors compared to case-matched normal tissues (n = 119, both P < 0.0001), and correlate with markedly reduced survival (P = 0.007 and P = 0.041, respectively). Furthermore, CDK2 and PAF are also associated with poor prognosis in certain subtypes of breast cancer (n = 1802) and gastric cancer (n = 233). Although CDK2 could not significantly mediate the biological function of miR-200b, PAF siRNA knockdown phenocopied while restored expression of PAF abrogated the biological effects of miR-200b on ESCC cells. Moreover, PAF was revealed to mediate the inhibitory effects of miR-200b on Wnt/ß-Catenin signaling. Collectively, the pleiotropic effects of miR-200b in ESCC highlight its potential for therapeutic intervention in this aggressive disease.

RGD peptide targeted lipid-coated nanoparticles for combinatorial delivery of sorafenib and quercetin against hepatocellular carcinoma.

CONTEXT: Combination therapies provide a potential solution to address the tumor heterogeneity and drug resistance issues by taking advantage of distinct mechanisms of action of the multiple therapeutics. OBJECTIVE: To design arginine-glycineaspartic acid (RGD) modified lipid-coated nanoparticles (NPs) for the co-delivery of the hydrophobic drugs against hepatocellular carcinoma (HCC). MATERIALS AND METHODS: RGD modified lipid-coated PLGA NPs were developed for the targeted delivery of both sorafenib (SRF) and quercetin (QT) (RGD-SRF-QT NPs). Chemical-physical characteristics and release profiles were evaluated. In vitro cell viability assays were carried out on HCC cells. In vivo antitumor efficacies were evaluated in HCC animal model. RESULTS AND DISCUSSION: The combination of SRF and QT formulations was more effective than the single drug formulations in both NPs and solution groups. RGD-SRF-QT NPs achieved the most significant tumor growth inhibition effect in vitro and in vivo. CONCLUSION: The resulting NPs could provide a promising platform for co-delivery of multiple anticancer drugs for achievement of combinational therapy and could offer potential for enhancing the therapeutic efficacy on HCC.

The PI3K/AKT/c-MYC Axis Promotes the Acquisition of Cancer Stem-Like Features in Esophageal Squamous Cell Carcinoma.

The importance of intratumoral heterogeneity has been highlighted by the identification and characterization of cancer stem cells (CSCs). Based on the differential responsiveness to a Sox2 reporter, SRR2, we had found a novel dichotomy in esophageal squamous cell carcinoma (ESCC) cells, with reporter-responsive (RR) cells showing more CSC-like features than reporter-unresponsive (RU) cells. Specifically, RR cells exhibited significantly higher tumorsphere formation capacity, proportions of CD44(High) cells, chemoresistance to cisplatin, and tumorigenic potential in vivo. H2 O2 , a potent inducer of oxidative stress and reactive oxygen species, was found to induce a conversion from RU to RR cells; importantly, converted RR cells acquired CSC-like features. The PI3K/AKT/c-MYC signalling axis is important in this context, since pharmacologic blockade of PI3K-AKT or siRNA knockdown of c-MYC effectively inhibited the RR phenotype and its associated CSC-like features, as well as the H2 O2 -induced RU/RR conversion. In a cohort of 188 ESCC patient samples, we found a significant correlation between strong c-MYC expression and a short overall survival (p = .009). In conclusion, we have described a novel intratumoral heterogeneity in ESCC. The identification of the PI3K/AKT/c-MYC axis as a driver of CSC-like features carries therapeutic implications. Stem Cells 2016;34:2040-2051.

Silibinin suppresses NPM-ALK, potently induces apoptosis and enhances chemosensitivity in ALK-positive anaplastic large cell lymphoma.

Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK), an oncogenic fusion protein carrying constitutively active tyrosine kinase, is known to be central to the pathogenesis of ALK-positive anaplastic large cell lymphoma (ALK+ALCL). Here, it is reported that silibinin, a non-toxic naturally-occurring compound, potently suppressed NPM-ALK and effectively inhibited the growth and soft agar colony formation of ALK+ALCL cells. By western blots, it was found that silibinin efficiently suppressed the phosphorylation/activation of NPM-ALK and its key substrates/downstream mediators (including STAT3, MEK/ERK and Akt) in a time- and dose-dependent manner. Correlating with these observations, silibinin suppressed the expression of Bcl-2, survivin and JunB, all of which are found to be upregulated by NPM-ALK and pathogenetically important in ALK+ALCL. Lastly, silibinin augmented the chemosensitivity of ALK+ALCL cells to doxorubicin, particularly the small cell sub-set expressing the transcriptional activity of Sox2, an embryonic stem cell marker. To conclude, the findings suggest that silibinin might be useful in treating ALK+ALCL.

The Opposing Function of STAT3 as an Oncoprotein and Tumor Suppressor Is Dictated by the Expression Status of STAT3ß in Esophageal Squamous Cell Carcinoma.

PURPOSE: STAT3 is known to have both oncogenic and tumor suppressive effects, but the regulation of these opposing effects is elusive. We hypothesized that STAT3ß, one of the two STAT3 isoforms, is the key determinant in this context. EXPERIMENTAL DESIGN: The prognostic significance of STAT3ß and phospho-STAT3α(Y705) (pSTAT3α(Y705)) was evaluated in 286 cases of patients with esophageal squamous cell carcinoma (ESCC). STAT3ß-induced changes in the chemosensitivity to cisplatin and 5-fluorouracil were assessed both in vitro and in vivo. STAT3ß-induced changes in the frequency of cancer stem cells were evaluated using Hoechst and CD44 staining. How STAT3ß regulates STAT3α was determined using immunoprecipitation, confocal microscopy, DNA-binding, and chromatin immunoprecipitation-PCR. RESULTS: STAT3ß expression is an independent protective prognostic marker in patients with ESCC, which strongly correlated with longer overall survival (P = 0.0009) and recurrence-free survival (P = 0.0001). STAT3ß significantly decreased the cancer stem cell population, and sensitized ESCC cells to cisplatin and 5-fluorouracil in tumor xenografts. Mechanistically, STAT3ß markedly attenuated the transcription activity of STAT3α via inducing STAT3α:STAT3ß heterodimers. However, the heterodimer formation decreased the binding between STAT3α and PTPN9 (better known as PTP-MEG2), a protein tyrosine phosphatase, thereby promoting the phosphorylation of STAT3α(Y705) and enhancing its nuclear translocation and DNA binding. Correlating with this, high STAT3ß expression converts the prognostic value of pSTAT3α(Y705) from unfavorable to favorable in patients with ESCC. CONCLUSIONS: STAT3ß suppresses chemoresistance and cancer stemness by blocking the transcriptional activity of STAT3α. The paradoxical increase in pSTAT3α(Y705) induced by STAT3ß carries important implications as to how the biologic and prognostic significance of STAT3 in cancers should be interpreted.

Loss of miR-200b promotes invasion via activating the Kindlin-2/integrin ß1/AKT pathway in esophageal squamous cell carcinoma: An E-cadherin-independent mechanism.

Our previous studies have shown that loss of miR-200b enhances the invasiveness of esophageal squamous cell carcinoma (ESCC) cells. However, whether the miR-200-ZEB1/2-E-cadherin regulatory cascade, a master regulator of epithelial-to-mesenchymal transition (EMT), is involved in the regulation of ESCC invasion remains elusive. Here, we show that miR-200b represses ESCC cell invasion in vivo without altering the expression of E-cadherin and vimentin, two surrogate markers of EMT. However, an inverse correlation was observed between the expression levels of miR-200b and ZEB1/2 in both ESCC cell lines (n = 7, P < 0.05) and ESCC tumor samples (n = 88, P < 0.05). Methylation of E-cadherin gene was found to block the regulation of E-cadherin by the miR-200b-ZEB1/2 axis, indicating that an E-cadherin-independent mechanism can mediate the biological function of miR-200b in ESCC. We revealed that miR-200b suppresses the integrin ß1-AKT pathway via targeting Kindlin-2 to mitigate ESCC cell invasiveness. In two independent cohorts of ESCC samples (n = 20 and n = 53, respectively), Kindlin-2 expression positively correlated with the activation status of both the integrin signaling pathway and the PI3K-AKT signaling pathway (both P < 0.01). These data highlight that suppression of the Kindlin-2-integrin ß1-AKT regulatory axis is an alternative mechanism underlying the tumor suppressor function of miR-200b in ESCC.