c-Met Signaling Pathway Research Articles

Research Progress of Small Molecule VEGFR/c-Met Inhibitors as Anticancer Agents (2016-Present).

Vascular endothelial growth factor receptor 2 (VEGFR-2) binds to VEGFR-A, VEGFR-C and VEGFR-D and participates in the formation of tumor blood vessels, mediates the proliferation of endothelial cells, enhances microvascular permeability, and blocks apoptosis. Blocking or downregulating the signal transduction of VEGFR is the main way to discover new drugs for many human angiogenesis-dependent malignancies. Mesenchymal epithelial transfer factor tyrosine kinase (c-Met) is a high affinity receptor for hepatocyte growth factor (HGF). Abnormal c-Met signaling plays an important role in the formation, invasion and metastasis of human tumors. Therefore, the HGF/c-Met signaling pathway has become a significant target for cancer treatment. Related studies have shown that the conduction of the VEGFR and c-Met signaling pathways has a synergistic effect in inducing angiogenesis and inhibiting tumor growth. In recent years, multi-target small molecule inhibitors have become a research hotspot, among which the research of VEGFR and c-Met dual-target small molecule inhibitors has become more and more extensive. In this review, we comprehensively summarize the chemical structures and biological characteristics of novel VEGFR/c-Met dual-target small-molecule inhibitors in the past five years.

Anticancer and Antiangiogenic Activities of Novel α-Mangostin Glycosides in Human Hepatocellular Carcinoma Cells via Downregulation of c-Met and HIF-1α.

Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and is a leading cause of cancer-related death worldwide. Therefore, exploring effective anticancer agents and their modes of action is essential for the prevention and treatment of HCC. Glycosylation can significantly improve the physicochemical and biological properties of small molecules, such as high solubility, stability increase, and lower toxicity. In the present study, for the first time, we evaluated the anticancer and antiangiogenic activities of α-mangostin-3-O-β-D-2-deoxyglucopyranoside (Man-3DG) and α-mangostin 6-O-β-D-2-deoxyglucopyranoside (Man-6DG), glycosides of α-mangostin, against human HCC cells. Our results demonstrated that Man-3DG and Man-6DG significantly suppressed the growth of three different HCC cells (Hep3B, Huh7, and HepG2) as well as the migration of Hep3B cells. Furthermore, they induced cell cycle arrest in the G0/G1 phases and apoptotic cell death by regulating apoptosis-related proteins of mitochondria in Hep3B cells. Noticeably, Man-3DG and Man-6DG also caused autophagy, while co-treatment of the α-mangostin glycosides with an autophagy inhibitor 3-MA enhanced the inhibitory effect on Hep3B cell growth in comparison to single agent treatment. Moreover, Man-3DG and Man-6DG inhibited the c-Met signaling pathway that plays a critical role in the pathogenesis of HCC. Furthermore, the α-mangostin glycosides decreased Hep3B cell-induced angiogenesis in vitro through the downregulation of hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF). Notably, Man-6DG more effectively inhibited the growth, tumorsphere formation, and expression of cancer stemness regulators compared to α-mangostin and Man-3DG in 3D spheroid-cultured Hep3B cells. These findings suggest that the α-mangostin glycosides might be promising anticancer agents for HCC treatment with superior pharmacological properties than the parent molecule α-mangostin.

Text mining in a literature review of urothelial cancer using topic model

BackgroundUrothelial cancer (UC) includes carcinomas of the bladder, ureters, and renal pelvis. New treatments and biomarkers of UC emerged in this decade. To identify the key information in a vast amount of literature can be challenging. In this study, we use text mining to explore UC publications to identify important information that may lead to new research directions.MethodWe used topic modeling to analyze the titles and abstracts of 29,883 articles of UC from Pubmed, Web of Science, and Embase in Mar 2020. We applied latent Dirichlet allocation modeling to extract 15 topics and conducted trend analysis. Gene ontology term enrichment analysis and Kyoto encyclopedia of genes and genomes pathway analysis were performed to identify UC related pathways.ResultsThere was a growing trend regarding UC treatment especially immune checkpoint therapy but not the staging of UC. The risk factors of UC carried in different countries such as cigarette smoking in the United State and aristolochic acid in Taiwan and China. GMCSF, IL-5, Syndecan-1, ErbB receptor, integrin, c-Met, and TRAIL signaling pathways are the most relevant biological pathway associated with UC.ConclusionsThe risk factors of UC may be dependent on the countries and GMCSF, IL-5, Syndecan-1, ErbB receptor, integrin, c-Met, and TRAIL signaling pathways are the most relevant biological pathway associated with UC. These findings may provide further UC research directions.

Test identifies ovarian cancer patients with hyperactive c-Met and ErbB signaling tumors who may benefit from c-Met and pan-HER combination therapy.

e18038 Background: No sub-groups of ovarian cancer patients with clinically actionable ErbB genetic variants have yet been found. Measurement of ErbB signaling activity rather than genetic variants may identify ovarian cancer patients likely to benefit from ErbB targeted therapies. We have previously reported studies detecting dysregulated ErbB and c-Met signaling activity in 20%-25% of HER2-negative breast cancer patient tumors using the CELx Signaling Function Test. To determine whether ErbB or c-Met signaling dysregulation is involved in ovarian cancer, the CELx Test was adapted to analyze ovarian tumor cells. The current study set out to: 1) characterize c-Met and ErbB family signaling activity in ovarian patient tumor cells and tumor cell lines; 2) evaluate in vivo response to pan-HER and c-Met inhibitors in an ovarian tumor xenograft model. Methods: For the ex vivo studies, a set of fresh tumor specimens from 15 ovarian cancer patients and a set of 12 ovarian cancer cell lines was obtained. Cell samples were cultured from each. Live cell response to ErbB and c-Met agonists (NRG1b, EGF, or HGF) with or without an antagonist (2C4, a HER2 dimerization inhibitor or tepotinib, a c-Met kinase inhibitor) was measured using an xCELLigence impedance biosensor (Agilent Technologies). Signaling activity above a previously established cut-off value was used to identify abnormal levels of EGFR, HER2 and c-Met signaling activity. For the xenograft study, OVCAR-4, an ovarian cancer cell line, determined to have abnormal HER2, EGFR, and c-Met signaling with the CELx Test, was studied. 40 female NSG mice were each injected with two million cells. Mice were randomly assigned to either a control group or a treatment group that received neratinib, tepotinib, or neratinib and tepotinib for 16 days. Results: Of the patient cell and cell lines samples tested ex vivo with the CELx Test, 4 of 27 (15%; 95% CI = 6%-32%) had hyperactive HER2 and c-Met signaling pathways. In the xenograft study, tumor volume change was 40% less in the tepotinib + neratinib treated group than in the control arm change. There was no significant difference in tumor volume between the control and tepotinib or neratinib groups. Conclusions: These findings suggest that a significant sub-group of ovarian cancer patients have abnormal ErbB and c-Met signaling activity that may respond to treatment with a combination of ErbB and c-Met inhibitors. A clinical trial to evaluate treatment response of this patient sub-set to combined c-Met and pan-HER inhibitors is warranted.

Tivantinib inhibits the VEGF signaling pathway and induces apoptosis in gastric cancer cells with c-MET or VEGFA amplification.

Tivantinib has been described as a selective inhibitor of c-Met and is being studied in various types of cancer. In this study, we evaluated the effects of tivantinib on the suppression of gastric cancer (GC) cell migration and apoptosis. We also examined the mechanism of action of tivantinib by oncogenic pathway analysis. We applied an RNA-sequencing approach in 34 GC patients to identify oncogenes that are differentially expressed in GC tissues. To examine the inhibitory effect of tivantinib on GC cells, we conducted apoptosis analysis using an annexin V-APC/PI apoptosis detection kit and trans-well migration assay with human GC cell lines. For oncogenic pathway analysis, Western blot and quantitative real-time PCR analysis were used to detect the expression of proteins and genes before and after tivantinib exposure. In the RNA-sequencing analysis of 34 GC patients, c-Met and VEGFA genes were expressed and positively correlated with each other. Cell migration and apoptosis analysis demonstrated that tivantinib induced the best inhibition effect in SNU620, MKN45 (carries VEGFB mutation), AGS, and MKN28 cells, but not in KATO III (carries VEGFB and VEGFC mutations) cells. Oncogenic pathway analysis showed that tivantinib, in addition to c-Met signaling pathway inhibition, also inhibits VEGF signaling and MYC expression in VEGFA-expressing GC cells. We found that tivantinib has anti-cancer activity not only in GC cells overexpressing c-Met but also in non-c-Met GC cells by inhibition of the VEGF signaling pathway.

MP17-19 RECRUITED T CELLS PROMOTE THE BLADDER CANCER METASTASIS VIA UP-REGULATION OF THE ESTROGEN RECEPTOR β/IL-1/C-MET SIGNALS

You have accessJournal of UrologyBladder Cancer: Basic Research & Pathophysiology II (MP17)1 Apr 2020MP17-19 RECRUITED T CELLS PROMOTE THE BLADDER CANCER METASTASIS VIA UP-REGULATION OF THE ESTROGEN RECEPTOR β/IL-1/C-MET SIGNALS Le Tao, Jialin Meng*, Edward M. Messing, Jean V. Joseph, Chawnshang Chang, Jianxin Qiu, and ShuYuan Yeh Le TaoLe Tao More articles by this author , Jialin Meng*Jialin Meng* More articles by this author , Edward M. MessingEdward M. Messing More articles by this author , Jean V. JosephJean V. Joseph More articles by this author , Chawnshang ChangChawnshang Chang More articles by this author , Jianxin QiuJianxin Qiu More articles by this author , and ShuYuan YehShuYuan Yeh More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000842.020AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Bladder cancer (BCa) is the fourth most commonly occurring cancer in men and is the seventh leading cause of death from cancer among men and the eleventh among women in the United States. Previous studies have shown that higher ERβ expression rates in BCa are positively correlated to higher grades and stages, as well as muscle invasiveness, and that both ERα and ERβ signals affect BCa development. Immune cells may infiltrate into different types of cancers and influence cancer progression. An early study demonstrated that CD4+ T cells could promote prostate cancer invasion. In BCa, one report indicated that inflammation elicited from the tumor microenvironment (TME) may be able to influence tumor progression. METHODS: Tissue samples were obtained from 20 patients who were diagnosed with BCa. ERβ (PLKO.1-puro-shERβ) was constructed with the target sequence 5′ GCGAGTAACAAGGGCATGGAA-3′ into lentiviral pLKO.1 vector as ERβ sh-RNA. Chromatin immuno-precipitation (ChIP) assays were used to calculate the binding of ERβ-DNA complex. Female nude mice (6–8 week old) were purchased from NCI and implanted with 5637 cells that were engineered to express a luciferase reporter gene (lentiviral-luciferase). Images were detected 20 min after intraperitoneal injection of luciferin using a fluorescent imager (IVIS Spectrum, Caliper Life Sciences) at 5 different time points (3, 4, 5, 6 and 7 weeks after tumor cells implantation). RESULTS: Results from an in vitroco-culture system found that BCa recruited more CD4+ T cells than did normal bladder cells. The recruiting of T cells to BCa tissues may increase the proliferation and invasion of BCa cells. Mechanistic studies revealed that infiltrating T cells stimulate BCa estrogen receptor beta (ERβ) signaling and consequently increase the expression of the MET proto-oncogene, receptor tyrosine kinase (c-MET), through either direct binding to its promoter or viamodulation of IL-1 expression. Interruption of ERβ/c-MET or ERβ/IL-1/c-MET signaling viaERβ-shRNA, IL-1 antagonist, or the c-MET inhibitor, SU11274, could partially reverse the T cell-enhanced BCa cell invasion and proliferation. Finally, the mouse BCa model with xenografted BCa 5637 cells with T (HH) cells confirmed the results of in vitroco-culture studies. CONCLUSIONS: Infiltrating T cells could promote BCa metastasis viamodulation of the ERβ/c-MET or ERβ/IL-1/c-MET signaling pathways. Source of Funding: This work was partially supported by the University of Rochester CTSI (UL1 TR000042), and the National Natural Science Foundation of China. © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e233-e233 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Le Tao More articles by this author Jialin Meng* More articles by this author Edward M. Messing More articles by this author Jean V. Joseph More articles by this author Chawnshang Chang More articles by this author Jianxin Qiu More articles by this author ShuYuan Yeh More articles by this author Expand All Advertisement Loading ...

Targeting c-Met on gastric cancer cells through a fully human fab antibody isolated from a large naive phage antibody library.

The aberrant Hepatocyte growth factor (HGF)/ mesenchymal-epithelial transition factor (c-Met) signaling pathway in various malignancies and its correlation with tumor invasion and poor prognosis has validated c-Met as a compelling therapeutic target. Up to now, several monoclonal antibodies and small molecule inhibitors targeting c-Met have been introduced with different outcomes, none are yet clinically approved. Toward the generation of novel fully human anti-c-Met molecules, we generated a large naïve Fab antibody library using phage display technology, which subsequently screened for novel Fabs against c-Met. A phage library, with a functional size of 5.5 × 1010 individual antibody clones, was prepared using standard protocols and screened for c-Met-specific Fabs by successive rounds of panning. A panel of Fabs targeting c-Met were isolated, from which four clones were selected and further characterized by DNA sequencing. The c-Met binding ability of our selected Fabs was evaluated by c-Met ELISA assay and flow cytometry techniques. Among the confirmed anti-c-Met Fabs, clone C16, showed the highest affinity (Kaff: 0.3 × 109M-1), and 63% binding to MKN45 cells (a human gastric adenocarcinoma cell-line) as compared to c-Met negative T47D cell-line (9.03%). Together, our study presents a single-pot antibody library, as a valuable source for finding a range of antigen-specific Fab antibodies, and also, a fully human, high affinity and specific anti c-Met Fab antibody, C16, which has the potential of developing as a therapeutic or chemotherapeutic delivery agent for killing c-Met-positive tumor cells.

Light-Induced Radiosynthesis of 89Zr-DFO-Azepin-Onartuzumab for Imaging the Hepatocyte Growth Factor Receptor.

Methods that provide rapid access to radiolabeled antibodies are vital in the development of diagnostic and radiotherapeutic agents for PET or radioimmunotherapy. The human hepatocyte growth factor receptor (c-MET) signaling pathway is dysregulated in several malignancies, including gastric cancer, and is an important biomarker in drug discovery. Here, we used a photoradiochemical approach to produce 89Zr-radiolabeled onartuzumab (a monovalent, antihuman c-MET antibody), starting directly from the fully formulated drug (MetMAb). Methods: Simultaneous 89Zr-radiolabeling and protein conjugation was performed in one-pot reactions containing 89Zr-oxalate, the photoactive chelate desferrioxamine B (DFO)-aryl azide (DFO-ArN3), and MetMAb to give 89Zr-DFO-azepin-onartuzumab. As a control, 89Zr-DFO-benzyl Bn-isothiocyanate Bn-NCS-onartuzumab was prepared via a conventional two-step process using prepurified onartuzumab and DFO-Bn-NCS. Radiotracers were purified by using size-exclusion methods and evaluated by radiochromatography. Radiochemical stability was studied in human serum, and immunoreactivity was determined by cellular binding assays using MKN-45 gastric carcinoma cells. PET imaging at multiple time points (0-72 h) was performed on female athymic nude mice bearing subcutaneous MKN-45 xenografts. Biodistribution experiments were performed after the final image was obtained. The tumor specificity of 89Zr-DFO-azepin-onartuzumab was assessed in vivo by competitive inhibition (blocking) studies. Results: Initial photoradiosynthesis experiments produced 89Zr-DFO-azepin-onartuzumab in less than 15 min, with an isolated decay-corrected radiochemical yield (RCY) of 24.8%, a radiochemical purity of approximately 90%, and a molar activity of approximately 1.5 MBq nmol-1 Reaction optimization improved the radiochemical conversion of 89Zr-DFO-azepin-onartuzumab to 56.9% ± 4.1% (n = 3), with isolated RCYs of 41.2% ± 10.6% (n = 3) and radiochemical purity of more than 90%. Conventional methods produced 89Zr-DFO-Bn-NCS-onartuzumab with an isolated RCY of more than 97%, radiochemical purity of more than 97% and molar activity of approximately 14.0 MBq nmol-1 Both radiotracers were immunoreactive and stable in human serum. PET imaging and biodistribution studies showed high tumor uptake for both radiotracers. By 72 h, tumor and liver uptake (percentage injected dose [%ID]) reached 15.37 ± 5.21 %ID g-1 and 6.56 ± 4.03 %ID g-1, respectively, for 89Zr-DFO-azepin-onartuzumab (n = 4) and 21.38 ± 11.57 %ID g-1 and 18.84 ± 6.03 %ID g-1, respectively, for 89Zr-DFO-Bn-NCS-onartuzumab (n = 4). Blocking experiments gave a statistically significant reduction in tumor uptake (6.34 ± 0.47 %ID g-1) of 89Zr-DFO-azepin-onartuzumab (n = 4). Conclusion: The experiments demonstrated that photoradiosynthesis is a viable alternative approach for producing 89Zr-radiolabeled antibodies directly in protein formulation buffer, reducing protein aggregation and liver uptake.

Profiling of novel circulating microRNAs as a non-invasive biomarker in diagnosis and follow-up of high and low-grade gliomas

ObjectiveGlioblastoma (GBM) is the most common primary malignant neoplasm of the central nervous system (CNS). Despite the progress in therapeutic strategies such as surgical techniques, radiotherapy, chemotherapy, and targeted therapy, prognosis and therapeutically convenient monitoring tools in patients with GBM has not improved significantly up to now.Therefore, exosomal miRNAs as novel non-invasive biomarkers having high sensitivity and specificity are required to improve diagnosis and to develop new targeted therapy strategies for GBM patients. The aim of the present study was to investigate a novel miRNA signature as a predictive biomarker for diagnosis and measurement of response to therapeutic interventions in plasma of GBM patients versus traumatic brain injury and diffuse low-grade astrocytoma (LGA) patients. Patients and methodsPlasma exosomal-microRNAs were isolated from GBM (n = 25), LGA (n = 25), and head trauma patients (n = 15) as non-glioma control from March 2017 to June 2018 in Department of Neurosurgery at Rasoul-e-Akram Hospital. Through a bioinformatics analysis, we used Miranda, TargetScan, mirBase, DIANA-microT-CDS, and KEGG database as well as microarray data analysis from GEO for microRNA candidates. Finally, miR-210, miR-185, miR-5194, and miR-449 were selected among those miRNAs because they were recorded to target the maximum number of genes in EGFR and c-MET signaling pathways. Then, exosomal microRNAs were extracted from plasma of patients and quantitated by locked nucleic acid real-time PCR in GBM, LGA, and trauma patients. ResultsThis result is the first report on the role of circulating miR-185, miR-449, and miR-5194 in GBM compared to LGA and trauma. The plasma expression of miR-210 as an oncogenic miR was upregulated in GBM and LGA groups (P < 0.0001). Otherwise, miR-185, miR-5194, and miR-449 were significantly downregulated (P ≤ 0.05) in GBM and LGA compared to trauma patients. There was no significant downregulation in the expression of miR-185 between GBM and LGA, while the expression of miR-5194 (P ≤ 0.05) and miR-449 (P ≤ 0.05) was significantly decreased in GBM patients compared with LGA. ConclusionsThese results indicate that the levels of miR-210, miR-449, and miR-5194 are a promising diagnostic and prognostic biomarker positively correlated with histopathological grade and invasiveness of GBM. These findings imply that circulating microRNA can be potentially used as novel biomarkers for glioma that might be beneficial in clinical management of glioma patients.

Augmenting nab-paclitaxel/gemcitabine standard chemotherapy response by merestinib, an inhibitor of c-MET, Axl and DDR signaling pathways, in preclinical pancreatic cancer models

Background and Hypothesis: Pancreatic ductal adenocarcinoma (PDAC) is among the most lethal malignancies in Western countries. Nab-paclitaxel (NPT) plus gemcitabine (Gem) is the standard of care for PDAC leading to a dismal 8.5 months median survival. Aberrant signaling of c-MET, Axl and DDR have been reported in a variety of human cancers including PDAC. Merestinib (Mer) is a potent, smallmolecule inhibitor of these pathways. We evaluated the therapeutic efficacy of merestinib to enhance the antitumor response of standard chemotherapy in preclinical models of PDAC.&#x0D; Project Methods: Cell proliferation of PDAC-associated cells (AsPC-1, PANC-1 and fibroblasts) were evaluated by colorimetric WST-1 assay. Protein expression was determined by Western blot analysis. Tumor progression studies were performed in NOD/SCID mice.&#x0D; Results: In vitro studies demonstrated that both nab-paclitaxel plus gemcitabine and merestinib suppressed cell proliferation of PDAC epithelial cells and stromal cells. Importantly, the combination treatment demonstrated additive inhibitory effects. In AsPC-1 cells, at the medium dose level, NPT+Gem, Mer and NPT+Gem+Mer treatments inhibited cell proliferation by 53.9%, 13.5%, and 81.61%, respectively. In PANC-1 cells, at the highest dose level, inhibition in cell proliferation by NPT+Gem, Mer and NPT+Gem+Mer treatments was 53.6%, 3.7%, and 72.8%. In the PDAC-associated fibroblasts, at the medium dose level, NPT+Gem, Mer and NPT+Gem+Mer treatments inhibited growth by 55.3%, 58.0%, and 91.6%. Immunoblot analysis revealed that merestinib caused a decrease in the PI-3K-AKT signaling proteins and an increase in apoptosisrelated proteins cleaved PARP-1 or cleaved caspase-3 in PDAC cells either alone or in combination with nab-paclitaxel and gemcitabine. In vivo study to evaluate tumor growth inhibition effects of merestinib in a subcutaneous PDAC xenograft is currently ongoing.&#x0D; Conclusion: The antitumor effect of standard chemotherapy regimen can be significantly enhanced by the cMET/Axl/DDR pathway inhibitor merestinib, which may lead to clinically relevant therapeutic strategy to increased survival in PDAC patients.

Antitumor Activity of DFX117 by Dual Inhibition of c-Met and PI3Kα in Non-Small Cell Lung Cancer.

Aberrant activation of hepatocyte growth factor (HGF)/c-Met signaling pathway caused by gene amplification or mutation plays an important role in tumorigenesis. Therefore, c-Met is considered as an attractive target for cancer therapy and c-Met inhibitors have been developed with great interests. However, cancers treated with c-Met inhibitors inevitably develop resistance commonly caused by the activation of PI3K/Akt signal transduction pathway. Therefore, the combination of c-Met and PI3Kα inhibitors showed synergistic activities, especially, in c-Met hyperactivated and PIK3CA-mutated cells. In our previous study, we rationally designed and synthesized DFX117(6-(5-(2,4-difluorophenylsulfonamido)-6-methoxypyridin-3-yl)-N-(2-morpholinoethyl) imidazo[1,2-a]pyridine-3-carboxamide) as a novel PI3Kα selective inhibitor. Herein, the antitumor activity and underlying mechanisms of DFX117 against non-small cell lung cancer (NSCLC) cells were evaluated in both in vitro and in vivo animal models. Concurrent targeted c-Met and PI3Kα by DFX117 dose-dependent inhibited the cell growth of H1975 cells (PIK3CA mutation and c-Met amplification) and A549 cells (KRAS mutation). DFX117 subsequently induced G0/G1 cell cycle arrest and apoptosis. These data highlight the significant potential of DFX117 as a feasible and efficacious agent for the treatment of NSCLC patients.

A novel miR-365-3p/EHF/keratin 16 axis promotes oral squamous cell carcinoma metastasis, cancer stemness and drug resistance via enhancing \u03b25-integrin/c-met signaling pathway

BackgroundTargeting the c-Met signaling pathway has become a therapeutic strategy in multiple types of cancer. We unveiled a novel c-Met regulating mechanism that could be applied as a modality for oral squamous cell carcinoma (OSCC) therapy.MethodsUpregulation of keratin 16 (KRT16) was found by comparing isogenic pairs of low and high invasive human OSCC lines via microarray analysis. OSCC cells with ectopic expression or silencing of KRT16 were used to scrutinize functional roles and associated molecular mechanisms.ResultsWe observed that high KRT16 expression significantly correlated with poorer pathological differentiation, advanced stages, increased lymph nodes metastasis, and decreased survival rate from several Taiwanese OSCC patient cohorts. We further revealed that miR-365-3p could target ETS homologous factor (EHF), a KRT16 transcription factor, to decrease migration, invasion, metastasis and chemoresistance in OSCC cells via inhibition of KRT16. Under confocal microscopic examination, c-Met was found possibly partially associates with KRT16 through β5-integrin. Colocalization of these three proteins may facilitate c-Met and β5-integrin–mediated signaling in OSCC cells. Depletion of KRT16 led to increased protein degradation of β5-integrin and c-Met through a lysosomal pathway leading to inhibition of their downstream Src/STAT3/FAK/ERK signaling in OSCC cells. Knockdown of KRT16 enhanced chemosensitivity of OSCC towards 5-fluorouracil (5-FU). Various combination of c-Met inhibitor (foretinib), protein tyrosine kinase inhibitor (genistein), β5-integrin antibody, and 5-FU markedly augmented cytotoxic effects in OSCC cells as well as tumor killing effects in vitro and in vivo.ConclusionsOur data indicate that targeting a novel miR-365-3p/EHF/KRT16/β5-integrin/c-Met signaling pathway could improve treatment efficacy in OSCC.

PIM1 kinase promotes cell proliferation, metastasis and tumor growth of lung adenocarcinoma by potentiating the c-MET signaling pathway

The proto-oncogene PIM1 plays essential roles in proliferation, survival, metastasis and drug resistance in hematopoietic and solid tumors. Although PIM1 has been shown to be associated with lymph node metastasis and poor prognosis in non-small cell lung cancer, its underlying molecular mechanisms in this context are still unclear. Here we show that PIM1 is frequently overexpressed in lung adenocarcinomas, and its expression level is associated with c-MET expression and poor clinical outcome. We further demonstrate that PIM1 may regulate c-MET expression via phosphorylation of eukaryotic translation initiation factor 4B (eIF4B) on S406. Depletion of PIM1 decreased cell proliferation, migration, invasion and colony formation in vitro, as well as reduced tumor growth in vivo. And these effects were partially abrogated by restoring of c-MET expression. Our study implicates a promising therapeutic approach in lung adenocarcinoma patients with PIM1 and c-MET overexpression.

A curcumin derivative hydrazinobenzoylcurcumin suppresses stem-like features of glioblastoma cells by targeting Ca2+ /calmodulin-dependent protein kinase II.

Glioblastoma multiforme (GBM) is the most aggressive and common type of human primary brain tumor. Glioblastoma stem-like cells (GSCs) have been proposed to contribute to tumor initiation, progression, recurrence, and therapeutic resistance of GBM. Therefore, targeting GSCs could be a promising strategy to treat this refractory cancer. Calmodulin (CaM), a major regulator of Ca2+ -dependent signaling, controls various cellular functions via interaction with multiple target proteins. Here, we investigated the anticancer effect of hydrazinobenzoylcurcumin (HBC), a Ca 2+ /CaM antagonist, against GSCs derived from U87MG and U373MG cells. HBC significantly inhibited not only the self-renewal capacity, such as cell growth and neurosphere formation but also the metastasis-promoting ability, such as migration and invasion of GSCs. HBC induced apoptosis of GSCs in a caspase-dependent manner. Notably, HBC repressed the phosphorylation of Ca 2+ /CaM-dependent protein kinase II (CaMKII), c-Met, and its downstream signal transduction mediators, thereby reducing the expression levels of GSC markers, such as CD133, Nanog, Sox2, and Oct4. In addition, the knockdown of CaMKIIγ remarkably decreased the cancer stem cell-like phenotypes as well as the expression of stemness markers by blocking c-Met signaling pathway in U87MG GSCs. These results suggest that HBC suppresses the stem-like features of GBM cells via downregulation of CaM/CaMKII/c-Met axis and therefore CaMKII may be a novel therapeutic target to eliminate GSCs.

Protective effects of total saponins of Panax japonicas on non-alcoholic fatty liver disease through regulating expression of miR-199-5p

To research the effection and probable mechanism for the total saponins of Panax japonicas(TPSJ) in mice on non-alcoholic fatty liver disease. Forty SPF male Kunming mice were randomily divided into four group:control group,NAFLD group, low-dose TPSJ treated group,high-dose TPSJ treated group. High-fatty and high-frutose-diet was applied to eatablish NAFLD model，and TPSJ (100 and 200 mg·kg⁻¹) in feeding were given for the TPSJ groups for 4 weeks. To collect the serum with liver and the ALT and TC of serum were monitored after 4 weeks. The hepatic histopathologic structure was observed by haematoxylin-eosin (HE) staining, RT-PCR and RT-qPCR was applied for the detection of miR-199-5p，VEGFa，HGF，c-Met and protein expression level was detected bv laser confocal microscope.Compared with control group, the level of serum ALT and TC in the model group was higher,the liver of the model group showed that hepatocytes display obvious lipid deposition. Then TPSJ treated showed that markedly improved histopathologic changes, decreased fatty deposition. In the meantime,the expression level of miR-199-5p was significantly decreased, thus the expression of HGF and c-Met were significantly increased. TPSJ play a role of prevention on fatty liver, the machanism maybe by blocking miR-199-5p targeted to c-Met signaling pathways in NAFLD.

Histone demethylase JARID1B/KDM5B promotes aggressiveness of non-small cell lung cancer and serves as a good prognostic predictor

BackgroundLung cancer is the leading cause of cancer death worldwide. Recently, epigenetic dysregulation has been known to promote tumor progression and therefore may be a therapeutic target for anticancer therapy. JARID1B, a member of histone demethylases, has been found to be related to tumorigenesis in certain kinds of cancers. However, its biological roles in non-small cell lung cancer (NSCLC) remain largely unclear.MethodsWe firstly examined the expression of JARID1B in surgical specimens and six NSCLC cell lines. Then, we evaluated the relationship between JARID1B expression and clinicopathologic parameters in 72 NSCLC patients, thereby established its prognostic importance. We subsequently studied the functional roles of JARID1B in tumorigenesis to verify its clinicopathologic significance.ResultsOur results showed that JARID1B was overexpressed in NSCLC cells and JARID1B overexpression was associated with tumor size, lymph node metastasis, advanced stages, and poor overall survival in NSCLC patients. JARID1B overexpression resulted in increased cell proliferation and formation of tumorspheres and correlated positively with the expression of cancer stem cells (CSCs) and epithelial-mesenchymal transition (EMT) markers, while the c-Met signaling pathway was actively involved. It also correlated with the strength of resistance to cisplatin and doxorubicin. On the contrary, downregulation of JARID1B expression by applying shRNA or JARID1B inhibitor PBIT reversed these phenomena.ConclusionsJARID1B worsens prognosis of NSCLC patients by promotion of tumor aggressiveness through multiple biological facets which were associated with activation of the c-Met signaling, and can be a novel prognostic biomarker and therapeutic target for NSCLC.

C-Met affects gemcitabine resistance during carcinogenesis in a mouse model of pancreatic cancer.

Pancreatic adenocarcinoma is thought to develop from histologically identifiable intraductal lesions known as pancreatic intraepithelial neoplasias (PanINs), which exhibit similar morphological and genetic features to pancreatic ductal adenocarcinoma (PDAC). Therefore, a better understanding of the biological features underlying the progression of PanIN is essential to development more effective therapeutic interventions for PDAC. In recent years, numerous studies have reported that MET proto-oncogene receptor tyrosine kinase (c-MET) is a potential marker of pancreatic cancer stem cells (CSCs). CSCs have been revealed to initiate and propagate tumors in vitro and in vivo, and are associated with a chemoresistant phenotype. However, in vivo models using a xenograft approach are limited. In the present study, the morphological phenotype, molecular alteration and biological behavior of neoplasia in Pdx-1Cre/+, KrasLSL-G12D/+ and Metflox/flox and wild-type mice was analyzed. The results demonstrated that while oncogenic KrasLSL-G12D/+ increased PanIN initiation and significantly decreased survival rate compared with wild-type mice, no additive effect of c-Met receptor signaling on PanIN progression or prognosis was observed. Following gemcitabine administration, c-Met inhibition in Kras LSL-G12D/+ mice significantly decreased the total surface area of PanIN lesions and the number of anti-proliferation marker protein Ki-67 positive cells occupying PanIN lesions compared with Met+/+ mice. In conclusion, complete inhibition of the c-Met signaling pathway with chemotherapy may be useful for the treatment of pancreatic cancer.

Development of a SPECT Tracer to Image c-Met Expression in a Xenograft Model of Non-Small Cell Lung Cancer.

Elevated expression of the c-Met receptor plays a crucial role in cancers. In non-small cell lung cancer (NSCLC), aberrant activation of the c-Met signaling pathway contributes to tumorigenesis and cancer progression and may mediate acquired resistance to epidermal growth factor receptor-targeted therapy. c-Met is therefore emerging as a promising therapeutic target for NSCLC, and methods for noninvasive in vivo assessment of c-Met expression would improve NSCLC treatment and diagnosis. Methods: We developed a new c-Met-binding peptide (cMBP) radiotracer, 99mTc-hydrazine nicotinamide (HYNIC)-cMBP, for SPECT imaging. Cell uptake assays were performed on 2 NSCLC cell lines with different c-Met expressions: H1993 (high expression) and H1299 (no expression). In vivo tumor specificity was assessed by SPECT imaging in tumor-bearing mice at 0.5, 1, 2, and 4 h after injection of the probe. Blocking assays, biodistribution, and autoradiography were also conducted to determine probe specificity. Results:99mTc-HYNIC-cMBP was prepared with high efficiency and showed higher uptake in H1993 cells than in H1299 cells. Biodistribution and autoradiography also showed significantly higher percentages of the injected dose for 99mTc-HYNIC-cMBP in H1993 tumors than in H1299 tumors at 0.5 h (4.74 ± 1.43%/g and 1.00 ± 0.37%/g, respectively; P < 0.05). H1993 tumors were clearly visualized at 0.5 h in SPECT images, whereas H1299 tumors were not observed at any time. The specificity of 99mTc-HYNIC-cMBP for c-Met was demonstrated by a competitive block with an excess of nonradiolabeled peptide. Conclusion: For c-Met-targeted SPECT imaging of NSCLC, we developed 99mTc-HYNIC-cMBP, a tracer that specifically binds to c-Met with favorable pharmacokinetics in vitro and in vivo.

Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma

Recent data suggest that glioblastomas (GBM) activate the c-MET signaling pathway and display increased levels in anti-apoptotic Bcl-2 family members. Therefore, targeting these two deregulated pathways for therapy might yield synergistic treatment responses. We applied extracellular flux analysis to assess tumor metabolism. We found that combined treatment with ABT263 and Crizotinib synergistically reduces the proliferation of glioblastoma cells, which was dependent on dual inhibition of Bcl-2 and Bcl-xL. The combination treatment led to enhanced apoptosis with loss of mitochondrial membrane potential and activation of caspases. On the molecular level, c-MET-inhibition results in significant energy deprivation with a reduction in oxidative phosphorylation, respiratory capacity and a suppression of intracellular energy production (ATP). In turn, loss of energy levels suppresses protein synthesis, causing a decline in anti-apoptotic Mcl-1 levels. Silencing of Mcl-1 enhanced ABT263 and MET-inhibitor mediated apoptosis, but marginally the combination treatment, indicating that Mcl-1 is the central factor for the induction of cell death induced by the combination treatment. Finally, combined treatment with BH3-mimetics and c-MET inhibitors results in significantly smaller tumors than each treatment alone in a PDX model system of glioblastoma. These results suggest that c-MET inhibition causes a selective vulnerability of GBM cells to Bcl-2/Bcl-xL inhibition.

Recruited T cells promote the bladder cancer metastasis via up-regulation of the estrogen receptor β/IL-1/c-MET signals

Clinical data indicates that T cells can be recruited to bladder cancer (BCa), yet the impact of T cells on BCa progression remains unclear. In the present study, we found that T cells were recruited more to BCa tissues than to the surrounding normal bladder tissues. Results from an in vitro co-culture system also found that BCa recruited more CD4+ T cells than did normal bladder cells. The recruiting of T cells to BCa tissues may increase the proliferation and invasion of BCa cells. Mechanistic studies revealed that infiltrating T cells stimulate BCa estrogen receptor beta (ERβ) signaling and consequently increase the expression of MET proto-oncogene, receptor tyrosine kinase (c-MET), through either direct binding to its promoter or via modulation of IL-1 expression. Interruption of ERβ/c-MET or ERβ/IL-1/c-MET signaling via ERβ-shRNA, IL-1 antagonist, or the c-MET inhibitor, SU11274, could partially reverse the T cell-enhanced BCa cell invasion and proliferation. Finally, the mouse BCa model with xenografted BCa 5637 cells with T (HH) cells confirmed the results of in vitro co-culture studies showing that infiltrating T cells could promote BCa metastasis via modulation of the ERβ/c-MET or ERβ/IL-1/c-MET signaling pathways. These findings may provide a new therapeutic approach to better combat BCa progression via targeting these newly identified signaling pathways.