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  • Correction to: IL-33 facilitates proliferation of colorectal cancer dependent on COX2/PGE2
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-17
    Yongkui Li; Jie Shi; Shanshan Qi; Jian Zhang; Dong Peng; Zhenzhen Chen; Guobin Wang; Zheng Wang; Lin Wang

    In the original publication of this manuscript [1], there are three errors in Fig. 1. The identified errors do not affect the conclusions of the work.

    更新日期:2020-01-17
  • Musashi2 promotes EGF-induced EMT in pancreatic cancer via ZEB1-ERK/MAPK signaling
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-17
    Weiwei Sheng; Xiaoyang Shi; Yiheng Lin; Jingtong Tang; Chao Jia; Rongxian Cao; Jian Sun; Guosen Wang; Lei Zhou; Ming Dong

    Our previous study showed Musashi2 (MSI2) promoted chemotherapy resistance and pernicious biology of pancreatic cancer (PC) by down-regulating Numb and p53. We further explored the novel molecular mechanism involving its oncogenic role in PC development. We investigated the potential role and mechanism of MSI2 in EGF-induced EMT in PC in vitro and vivo. EGF enhanced EGFR (epidermal growth factor receptor) phosphorylation, induced EMT and activated ZEB1-ERK/MAPK signaling in 2 PC cells. However, MSI2 silencing reversed EGF stimulated function, including inhibiting EGF-promoted EMT-like cell morphology and EGF-enhanced cell invasion and migration. Meanwhile, MSI2 silencing inhibited EGF-enhanced EGFR phosphorylation at tyrosine 1068 and reversed EGF-induced change of the key proteins in EMT and ZEB1-ERK/MAPK signaling (ZEB1, E-cad, ZO-1, β-catenin, pERK and c-Myc). Additionally, MSI2 was co-stained and co-immunoprecipitated with ZEB1, pERK and c-Myc in PC cells by IF and co-IP, implying a close interaction between them. In vivo, MSI2 silencing inhibited pancreatic tumor size in situ and distant liver metastases. A close relationship of MSI2 with EMT and ZEB1-ERK/MAPK signaling were also observed in vivo and human PC samples, which coordinately promoted the poor prognosis of PC patients. MSI2 promotes EGF-induced EMT in PC via ZEB1-ERK/MAPK signaling.

    更新日期:2020-01-17
  • De-regulated STAT5A/miR-202-5p/USP15/Caspase-6 regulatory axis suppresses CML cell apoptosis and contributes to Imatinib resistance
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-17
    Zi-Yuan Nie; Min Yao; Zhan Yang; Lin Yang; Xiao-Jun Liu; Jing Yu; Ying Ma; Nan Zhang; Xiao-Yan Zhang; Meng-Han Liu; Ling-Ling Jiang; Jian-Min Luo

    STAT5 plays an important role in the transformation of hematopoietic cells by BCR-ABL. However, the downstream target genes activated by STAT5 in chronic myeloid leukemia (CML) cells remain largely unclear. Here, we investigated the mechanistic functional relationship between STAT5A-regulated microRNA and CML cell apoptosis. The expression of USP15, Caspase-6, STAT5A-regulated miR-202-5p and STAT5A was detected by qRT-PCR and Western blotting in CML cell lines and PBMCs of CML patients. Cell apoptosis was evaluated by flow cytometry. Both gain- and loss-of-function experiments were used to investigate the roles of USP15, miR-202-5p and STAT5A in CML. Luciferase reporter assay detected the effect of miR-202-5p on USP15 expression. Xenograft animal model was used to test the effect of anti-miR-202-5p and pimozide on K562 cell xenograft growth. USP15 expression was significantly downregulated in CML cell lines and PBMCs of CML patients. Depletion of USP15 increased, whereas overexpression of USP15 reduced the resistance of CML cells to Imatinib. Further, decreased deubiquitinating activity of USP15 by USP15 downregulation led to reduced caspase-6 level, thus attenuating CML cell apoptosis. Mechanistically, miR-202-5p was upregulated in K562G cells and negatively regulated USP15 expression by directly targeting USP15 3′-UTR. Correspondingly, upregulation of miR-202-5p enhanced the resistance of CML cells to Imatinib by inhibiting cell apoptosis. Importantly, STAT5A was upregulated in CML cells and directly activated miR-202-5p transcription by binding to the pre-miR-202 promoter. Pimozide induced CML cell apoptosis and significantly reduced K562 cell xenograft growth in vivo by blocking STAT5A/miR-202-5p/USP15/Caspase-6 regulatory axis. we provide the first evidence that de-regulated STAT5A/miR-202-5p/USP15/Caspase-6 regulatory axis suppresses the apoptosis of CML cells, targeting this pathway might be a promising therapeutic approach for the treatment of CML.

    更新日期:2020-01-17
  • Down-regulated lncRNA SBF2-AS1 inhibits tumorigenesis and progression of breast cancer by sponging microRNA-143 and repressing RRS1
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-17
    Wenfei Xia; Yun Liu; Teng Cheng; Tao Xu; Menglu Dong; Xiaopeng Hu

    Recently, the roles of long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) in human diseases have been unveiled, this research was conducted to explore the impacts of lncRNA SET-binding factor 2-antisense RNA1 (SBF2-AS1), miR-143 and resistance to ralstonia solanacearum 1 (RRS1) on breast cancer (BC) development. The expression of SBF2-AS1, miR-143 and RRS1 in BC tissues, as well as in MDA-MB-231 and MCF-7 cell lines were assessed. Subsequently, the cells were transfected with miR-143 mimics or/and silenced or overexpressed SBF2-AS1 plasmids, and their negative controls. Then the proliferation, colony formation ability, cell cycle arrest, apoptosis, invasion and migration of the cells were assessed through gain- and loss-of-function experiments. Furthermore, the tumor growth, ki-67 expression and apoptosis in vivo were observed by subcutaneous tumorigenesis in nude mice. Binding relation between SBF2-AS1 and miR-143, and that between miR-143 and RRS1 were confirmed. SBF2-AS1 and RRS1 were amplified, while miR-143 was reduced in BC tissues and cells. Reduced SBF2-AS1 and elevated miR-143 could repress the proliferation, invasion and migration via restraining RRS1 expression. Moreover, knockdown of SBF2-AS1 up-regulated miR-143 to promote the apoptosis of BC cells by downregulating RRS1, resulting in a prohibitive effect on the tumorigenesis and progression of BC. Results of in vivo experiments indicated that the inhibited SBF2-AS1 and overexpressed miR-143 could restrict BC cell proliferation and promote apoptosis, and decelerate tumor growth in xenografts. We have discovered in this study that down-regulated SBF2-AS1 could inhibit tumorigenesis and progression of BC by up-regulation miR-143 and repressing RRS1, which provides basic therapeutic considerations for a novel target against BC.

    更新日期:2020-01-17
  • Correction to: ZNF326 promotes malignant phenotype of glioma by up-regulating HDAC7 expression and activating Wnt pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-16
    Xinmiao Yu; Minghao Wang; Jingjing Wu; Qiang Han; Xiupeng Zhang

    In the original publication of this manuscript [1], the author mislabeled the CTL group and ZNF326 group in Fig. 2-I,J (MTT result). The revised Fig. 2 is shown below.

    更新日期:2020-01-16
  • Lysine demethylase 2 (KDM2B) regulates hippo pathway via MOB1 to promote pancreatic ductal adenocarcinoma (PDAC) progression
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-15
    Ming Quan; Zhiqin Chen; Feng Jiao; Xiuying Xiao; Qing Xia; Jingde Chen; Qian Chao; Yandong Li; Yong Gao; Haiyan Yang; Liwei Wang; Jiujie Cui

    Mps1 binding protein (MOB1) is one of the core components of the mammalian Hippo pathway and plays important roles in cancer development. However, its expression, function and regulation in pancreatic ductal adenocarcinoma (PDAC) have not been revealed yet. The expression of MOB1 and lysine demethylase 2B (KDM2B) in PDAC and adjacent normal pancreas tissues were measured. Also, the underlying mechanisms of altered MOB1 expression and its impact on PDAC biology were investigated. We revealed for the first time that MOB1 was decreased expression in PDAC and was a statistically significant independent predictor of poor survival, and restored expression of MOB1 suppressed the proliferation, migration and invasion of PDAC cells. Further studies demonstrated that KDM2B directly bound to the promoter region of MOB1, and suppressed the promoter activity of MOB1 and transcriptionally inhibited the MOB1 expression. Furthermore, KDM2B regulated Hippo pathway and promoted PDAC proliferation, migration and invasion via MOB1. This study demonstrated the mechanism and roles of a novel KDM2B/MOB1/Hippo signaling in PDAC progression.

    更新日期:2020-01-15
  • KDM4B facilitates colorectal cancer growth and glucose metabolism by stimulating TRAF6-mediated AKT activation
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-13
    Haijie Li; Jingqin Lan; Guihua Wang; Kaixuan Guo; Caishun Han; Xiaolan Li; Junbo Hu; Zhixin Cao; Xuelai Luo

    Histone lysine demethylase 4B (KDM4B) has been implicated in various pathological processes and human diseases. Glucose metabolism is the main pattern of energy supply in cells and its dysfunction is closely related to tumorigenesis. Recent study shows that KDM4B protects against obesity and metabolic dysfunction. We realized the significant role of KDM4B in metabolism. However, the role of KDM4B in glucose metabolism remains unclear. Here, we sought to delineate the role and mechanism of KDM4B in glucose metabolism in colorectal cancer (CRC). We first analyzed the role of KDM4B in glucose uptake and CRC growth. We then investigated the consequences of KDM4B inhibition on the expression of GLUT1 and AKT signaling, also explored the underlying mechanism. Finally, we detected the mechanism in vivo and assessed the potential correlation between the expression of KDM4B and CRC prognosis. We found that KDM4B promoted glucose uptake and ATP production by regulating the expression of GLUT1 via the AKT signaling pathway. KDM4B could interact with TRAF6 and promote TRAF6-mediated ubiquitination of AKT for AKT activation. Furthermore, we demonstrated that KDM4B was overexpressed in CRC specimens and high level of KDM4B was associated with a poor survival rate in CRC patients. These findings reveal that KDM4B plays an important role in promoting CRC progression by enhancing glucose metabolism.

    更新日期:2020-01-14
  • Circular RNA circRHOBTB3 acts as a sponge for miR-654-3p inhibiting gastric cancer growth
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-13
    Guangxu Deng; Tingyu Mou; Jiayong He; Da Chen; Daojun Lv; Hao Liu; Jiang Yu; Shuang Wang; Guoxin Li

    Circular RNAs (circRNAs) have recently emerged as a new family of noncoding RNAs that are involved in the causation and progression of various cancers. However, the roles of circRNAs in the tumorigenesis of gastric cancer (GC) are still largely unknown. The expression profiles of circRNAs in GC were identified in open GEO database and were evaluated at the mRNA level in clinical GC samples compared with paired non-tumorous tissues. Kaplan-Meier survival curve was used to analyze the correlation of circRNA and patients’ prognosis. Subsequently, the circular structures of candidate circRNAs were validated by Sanger sequencing, divergent primer PCR, and RNase R treatments. Gain- and loss-of-function analyses were performed to evaluate the functional significance of it in GC initiation and progression. Dual-luciferase reporter and RNA pull-down assays were used to identify the microRNA (miRNA) sponge mechanism of circRNAs. The expression of circRHOBTB3 was lower in GC tissues and cell lines. Downregulation of circRHOBTB3 was significantly correlated with poor differentiation and unfavorable prognosis in patients with GC. Overexpression of circRHOBTB3 in GC cells led to decreased proliferation and induced G1/S arrest in vitro, accompanied with inhibited xenograft tumor growth in vivo, while the opposite effects were achieved in circRHOBTB3-silenced cells. Furthermore, we demonstrated that circRHOBTB3 acts as a sponge for miR-654-3p and verified that p21 is a novel target of miR-654-3p. Taken together, this study revealed that circRHOBTB3 might function as competing endogenous RNA (ceRNA) for miR-654-3p, which could contribute to growth inhibition of GC through activating p21 signaling pathway. Our data suggested that circRHOBTB3 would serve as a novel promising diagnosis marker and therapeutic target for GC.

    更新日期:2020-01-13
  • Complement C3 overexpression activates JAK2/STAT3 pathway and correlates with gastric cancer progression
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-13
    Kaitao Yuan; Jinning Ye; Zhenguo Liu; Yufeng Ren; Weiling He; Jianbo Xu; Yulong He; Yujie Yuan

    Localized C3 deposition is a well-known factor of inflammation. However, its role in oncoprogression of gastric cancer (GC) remains obscured. This study aims to explore the prognostic value of C3 deposition and to elucidate the mechanism of C3-related oncoprogression for GC. From August to December 2013, 106 GC patients were prospectively included. The regional expression of C3 and other effectors in gastric tissues were detected by WB, IHC, qRT-PCR and other tests. The correlation of localized C3 deposition and oncologic outcomes was determined by 5-year survival significance. Human GC and normal epithelial cell lines were employed to detect a relationship between C3 and STAT3 signaling pathway in vitro experiments. C3 and C3a expression were markedly enhanced in GC tissues at both mRNA and protein levels compared with those in paired nontumorous tissues. According to IHC C3 score, 65 (61.3%) and 41 (38.7%) patients had high and low C3 deposition, respectively. C3 deposition was negatively correlated with plasma levels of C3 and C3a (both P < 0.001) and positively correlated with pathological T and TNM stages (both P < 0.001). High C3 deposition was identified as an independent prognostic factor of poor 5-year overall survival (P = 0.045). In vitro C3 administration remarkably enhanced p-JAK2/p-STAT3 expression in GC cell lines but caused a reduction of such activation when pre-incubated with a C3 blocker. Importantly, C3 failed to activate such signaling in cells pre-treated with a JAK2 inhibitor. Localized C3 deposition in the tumor microenvironment is a relevant immune signature for predicting prognosis of GC. It may aberrantly activate JAK2/STAT3 pathway allowing oncoprogression. ClinicalTrials.gov, NCT02425930, Registered 1st August 2013.

    更新日期:2020-01-13
  • Correction to: DNMT3b/OCT4 expression confers sorafenib resistance and poor prognosis of hepatocellular carcinoma through IL-6/STAT3 regulation
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-13
    Ssu-Chuan Lai; Yu-Ting Su; Ching-Chi Chi; Yung-Che Kuo; Kam-Fai Lee; Yu-Chih Wu; Pei-Chi Lan; Muh-Hwa Yang; Te-Sheng Chang; Yen-Hua Huang

    In the original publication of this article [1], labelling within Fig. 7a was incorrect. The updated figure is shown below, with ‘DMT1’ now corrected to read ‘DNMT1’.

    更新日期:2020-01-13
  • C3a-C3aR signaling promotes breast cancer lung metastasis via modulating carcinoma associated fibroblasts
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-13
    Chi Shu; Haoran Zha; Haixia Long; Xinxin Wang; Fei Yang; Jianbao Gao; Chunyan Hu; Li Zhou; Bo Guo; Bo Zhu

    Mounting evidence suggests that complement components promote tumor progression via modulating immune suppression, angiogenesis, or tumor cell proliferation. However, the role of C3a-C3aR signaling in regulating lung metastasis of breast cancer remains unknown. We performed various ex-vivo and in-vivo assays. Genetic and pharmacological C3aR blockade models were applied to investigate the role of C3a-C3aR in metastasis of breast cancer. C3a-C3aR signaling in CAFs facilitates the metastasis of breast cancer. Mechanically, C3a-C3aR signaling augments pro-metastatic cytokine secretion and extracellular matrix components expression of CAFs via the activation of PI3K-AKT signaling. Genetic or pharmacological blockade of C3aR signaling effectively inhibited lung metastasis of breast cancer in mouse models. C3a-C3aR signaling in CAFs facilitates the metastasis of breast cancer. Targeting C3aR signaling is a potential anti-metastasis strategy for breast cancer therapy.

    更新日期:2020-01-13
  • lncTUG1/miR-144-3p affect the radiosensitivity of esophageal squamous cell carcinoma by competitively regulating c-MET
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-09
    Pan Wang; Zhuanbo Yang; Ting Ye; Fei Shao; Jiagen Li; Nan Sun; Jie He

    Long noncoding RNAs (lncRNAs) are involved in the progression of various cancers and affect the response to radiotherapy. This study focused on clarifying the underlying mechanism by which lncTUG1 affects the radiosensitivity of esophageal squamous cell carcinoma (ESCC). lncTUG1, miR-144-3p and MET expression levels were detected in ESCC tissues and cells by qRT-PCR. Western blotting was used to examine the protein levels of MET, p-AKT and EGFR. The dual-luciferase reporter system and RNA immunoprecipitation (RIP) assays were used to confirm the interaction between lncTUG1 and miR-144-3p or miR-144-3p and MET. MTT, colony formation and flow cytometry assays were applied to examine the behavioral changes in EC9706 and KYSE30 cells. lncTUG1 was upregulated in ESCC cells and tissues, and lncTUG1 expression was associated with an advanced pathological stage. The bioinformatics analysis revealed that lncTUG1 could specifically bind to miR-144-3p, which was downregulated in ESCC. There was a negative correlation between lncTUG1 and miR-144-3p. LncTUG1 inhibition retarded proliferation and colony formation and induced apoptosis in ESCC cells. Moreover, lncTUG1 knockdown dramatically improved the effect of radiotherapy on ESCC development both in vivo and in vitro. Furthermore, MET was revealed as a downstream target of miR-144-3p and is downregulated by it. LncTUG1 promoted the progression of ESCC and elevated radiotherapy resistance in ESCC cells, accompanied by a high level of MET expression. Moreover, we found that knockdown of lncTUG1 enhanced the radiosensitivity of ESCC cells via the p-AKT signaling pathway. Our results indicate that lncTUG1 enhances the radiotherapy resistance of ESCC by lowering the miR-144-3p level and modulating the MET/EGFR/AKT axis.

    更新日期:2020-01-11
  • Targeting hypoxia in tumor: a new promising therapeutic strategy
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-10
    Maria Carla Bosco; Gabriella D’Orazi; Donatella Del Bufalo

    Low oxygen condition (hypoxia) is considered a hallmark of rapidly growing solid tumors. The presence of hypoxia renders tumor cells resistant to conventional chemo- and radio-therapy selecting a more malignant and invasive phenotype, and playing a negative role in patient prognosis. This commentary wishes to recognize the 2019 Nobel Prize in Medicine awarded to three physicians-scientists, Prof. William G. Kaelin Jr., Prof. Sir Peter J. Ratcliffe, and Prof. Gregg L. Semenza, for their discovery of the mechanisms mediating cell ability to sense and adapt to changes in oxygen availability. Their studies established the basis for our understanding of the role of hypoxia in a variety of diseases, including anemia, renal failure, cardiovascular disease, metabolic diseases, and cancer, paving the way for new promising therapeutic strategies through the development of drugs that can either activate or block the oxygen-sensing machinery.

    更新日期:2020-01-11
  • A pre-existing population of ZEB2+ quiescent cells with stemness and mesenchymal features dictate chemoresistance in colorectal cancer
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-08
    Federica Francescangeli; Paola Contavalli; Maria Laura De Angelis; Silvia Careccia; Michele Signore; Tobias Longin Haas; Federico Salaris; Marta Baiocchi; Alessandra Boe; Alessandro Giuliani; Olga Tcheremenskaia; Alfredo Pagliuca; Ombretta Guardiola; Gabriella Minchiotti; Lidia Colace; Antonio Ciardi; Vito D’Andrea; Filippo La Torre; JanPaul Medema; Ruggero De Maria; Ann Zeuner

    Quiescent/slow cycling cells have been identified in several tumors and correlated with therapy resistance. However, the features of chemoresistant populations and the molecular factors linking quiescence to chemoresistance are largely unknown. A population of chemoresistant quiescent/slow cycling cells was isolated through PKH26 staining (which allows to separate cells on the basis of their proliferation rate) from colorectal cancer (CRC) xenografts and subjected to global gene expression and pathway activation analyses. Factors expressed by the quiescent/slow cycling population were analyzed through lentiviral overexpression approaches for their ability to induce a dormant chemoresistant state both in vitro and in mouse xenografts. The correlation between quiescence-associated factors, CRC consensus molecular subtype and cancer prognosis was analyzed in large patient datasets. Untreated colorectal tumors contain a population of quiescent/slow cycling cells with stem cell features (quiescent cancer stem cells, QCSCs) characterized by a predetermined mesenchymal-like chemoresistant phenotype. QCSCs expressed increased levels of ZEB2, a transcription factor involved in stem cell plasticity and epithelial-mesenchymal transition (EMT), and of antiapototic factors pCRAF and pASK1. ZEB2 overexpression upregulated pCRAF/pASK1 levels resulting in increased chemoresistance, enrichment of cells with stemness/EMT traits and proliferative slowdown of tumor xenografts. In parallel, chemotherapy treatment of tumor xenografts induced the prevalence of QCSCs with a stemness/EMT phenotype and activation of the ZEB2/pCRAF/pASK1 axis, resulting in a chemotherapy-unresponsive state. In CRC patients, increased ZEB2 levels correlated with worse relapse-free survival and were strongly associated to the consensus molecular subtype 4 (CMS4) characterized by dismal prognosis, decreased proliferative rates and upregulation of EMT genes. These results show that chemotherapy-naive tumors contain a cell population characterized by a coordinated program of chemoresistance, quiescence, stemness and EMT. Such population becomes prevalent upon drug treatment and is responsible for chemotherapy resistance, thus representing a key target for more effective therapeutic approaches.

    更新日期:2020-01-08
  • Metformin-repressed miR-381-YAP-snail axis activity disrupts NSCLC growth and metastasis
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-06
    Dan Jin; Jiwei Guo; Yan Wu; Weiwei Chen; Jing Du; Lijuan Yang; Xiaohong Wang; Kaikai Gong; Juanjuan Dai; Shuang Miao; Xuelin Li; Guoming Su

    Recent evidence indicates that metformin inhibits mammalian cancer growth and metastasis through the regulation of microRNAs. Metformin regulates miR-381 stability, which plays a vital role in tumor progression. Moreover, increased YAP expression and activity induce non-small cell lung cancer (NSCLC) tumor growth and metastasis. However, the molecular mechanism underpinning how metformin-induced upregulation of miR-381 directly targets YAP or its interactions with the epithelial-mesenchymal transition (EMT) marker protein Snail in NSCLC is still unknown. Levels of RNA and protein were analyzed using qPCR, western blotting and immunofluorescence staining. Cellular proliferation was detected using a CCK8 assay. Cell migration and invasion were analyzed using wound healing and transwell assays. Promoter activity and transcription were investigated using the luciferase reporter assay. Chromatin immunoprecipitation was used to detect the binding of YAP to the promoter of Snail. The interaction between miR-381 and the 3′UTR of YAP mRNA was analyzed using the MS2 expression system and co-immunoprecipitation with biotin. We observed that miR-381 expression is negatively correlated with YAP expression and plays an opposite role to YAP in the regulation of cellular proliferation, invasion, migration, and EMT of NSCLC cells. The miR-381 function as a tumor suppressor was significantly downregulated in lung cancer tissue specimens and cell lines, which decreased the expression of its direct target YAP. In addition, metformin decreased cell growth, migration, invasion, and EMT via up-regulation of miR-381. Moreover, YAP, which functions as a co-transcription factor, enhanced NSCLC progression and metastasis by upregulation of Snail. Snail knockdown downregulated the mesenchymal marker vimentin and upregulated the epithelial marker E-cadherin in lung cancer cells. Furthermore, miR-381, YAP, and Snail constitute the miR-381-YAP-Snail signal axis, which is repressed by metformin, and enhances cancer cell invasiveness by directly regulating EMT. Metformin-induced repression of miR-381-YAP-Snail axis activity disrupts NSCLC growth and metastasis. Thus, we believe that the miR-381-YAP-Snail signal axis may be a suitable diagnostic marker and a potential therapeutic target for lung cancer.

    更新日期:2020-01-07
  • LncRNA LINC00662 promotes colon cancer tumor growth and metastasis by competitively binding with miR-340-5p to regulate CLDN8/IL22 co-expression and activating ERK signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-03
    Bo Cheng; Aimei Rong; Quanbo Zhou; Wenlu Li

    LncRNA LINC00662 is closely related to the occurrence and development of cancer. This study aims to explore the effect of LINC00662 on colon cancer tumor growth and metastasis and its molecular mechanism. CCK8, colony formation, transwell, scratch wound, TUNEL, flow cytometry, RT-PCR, western blotting and immunohistochemistry assays were used to detect the proliferation, apoptosis, invasion and migration of colon cancer cell and mRNA and protein expressions. Luciferase reporter and RNA pull down assays were used to detect the combination of LINC00662 and miR-340-5p or IL22 and the combination of miR-340-5p and CLDN8/IL22. Co-immunoprecipitation were used to detect the co-expression of CLDN8 and IL22 in colon cell lines. The targets of LINC00662 were predicated by Starbase v2.0. The target genes of miR-340-5p were predicated by miRDB and TargetScan. GO and KEGG enrichment analysis were performed by DAVID website. LINC00662 was up-regulation in colon cancer tissues and cell lines. Univariate Cox regression analysis showed that the LINC00662 expression level was related to the poor prognosis. LINC00662-WT and miR-340-5p mimics co-transfection depressed luciferase activity and IL22/CLDN8-WT and miR-340-5p inhibitors co-transfection memorably motivated luciferase activity. LINC00662 overexpression promoted cell proliferation, invasion and migration, and inhibited cell apoptosis in colon cancer. In vivo xenograft studies in nude mice manifested that LINC00662 overexpression prominently accelerate tumor growth. There was an opposite reaction in the biological functions of colon cells and tumor growth between LINC00662 overexpression and LINC00662 inhibition in vitro and in vivo. The functions of miR-340-5p mimics regulating the biological functions of colon cells and tumor growth were consistent with those of LINC00662 inhibition. CLDN8 and IL22, as target genes of miR-340-5p, reversed the functions of LINC00662 affecting the biological functions of colon cells and the protein levels of Bax, Bcl-2, XIAP, VEGF, MMP-2, E-cadherin and N-cadherin. Co-immunoprecipitation experiments indicated that CLDN8 directly interact with IL22 in colon cell lines. LINC00662 regulated CLDN8 and IL22 expressions and the activation of ERK signaling pathway via targeting miR-340-5p. LINC00662 overexpression promoted the occurrence and development of colon cancer by competitively binding with miR-340-5p to regulate CLDN8/IL22 co-expression and activating ERK signaling pathway.

    更新日期:2020-01-04
  • MiR-200c sensitizes Olaparib-resistant ovarian cancer cells by targeting Neuropilin 1
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-02
    Enrica Vescarelli; Giulia Gerini; Francesca Megiorni; Eleni Anastasiadou; Paola Pontecorvi; Luciana Solito; Claudia De Vitis; Simona Camero; Claudia Marchetti; Rita Mancini; Pierluigi Benedetti Panici; Carlo Dominici; Ferdinando Romano; Antonio Angeloni; Cinzia Marchese; Simona Ceccarelli

    Ovarian cancer (OC) is the most lethal gynecological malignancy and the second leading cause of cancer-related death in women. Treatment with PARP inhibitors (PARPi), such as Olaparib, has been recently introduced for OC patients, but resistance may occur and underlying mechanisms are still poorly understood. The aim of this study is to identify target genes within the tumor cells that might cause resistance to Olaparib. We focused on Neuropilin 1 (NRP1), a transmembrane receptor expressed in OC and correlated with poor survival, which has been also proposed as a key molecule in OC multidrug resistance. Using three OC cell lines (UWB, UWB-BRCA and SKOV3) as model systems, we evaluated the biological and molecular effects of Olaparib on OC cell growth, cell cycle, DNA damage and apoptosis/autophagy induction, through MTT and colony forming assays, flow cytometry, immunofluorescence and Western blot analyses. We evaluated NRP1 expression in OC specimens and cell lines by Western blot and qRT-PCR, and used RNA interference to selectively inhibit NRP1. To identify miR-200c as a regulator of NRP1, we used miRNA target prediction algorithms and Pearsons’ correlation analysis in biopsies from OC patients. Then, we used a stable transfection approach to overexpress miR-200c in Olaparib-resistant cells. We observed that NRP1 is expressed at high levels in resistant cells (SKOV3) and is upmodulated in partially sensitive cells (UWB-BRCA) upon prolonged Olaparib treatment, leading to poor drug response. Our results show that the selective inhibition of NRP1 is able to overcome Olaparib resistance in SKOV3 cells. Moreover, we demonstrated that miR-200c can target NRP1 in OC cells, causing its downmodulation, and that miR-200c overexpression is a valid approach to restore Olaparib sensitivity in OC resistant cells. These data demonstrate that miR-200c significantly enhanced the anti-cancer efficacy of Olaparib in drug-resistant OC cells. Thus, the combination of Olaparib with miRNA-based therapy may represent a promising treatment for drug resistant OC, and our data may help in designing novel precision medicine trials for optimizing the clinical use of PARPi.

    更新日期:2020-01-02
  • MiR-199a-modified exosomes from adipose tissue-derived mesenchymal stem cells improve hepatocellular carcinoma chemosensitivity through mTOR pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2020-01-02
    Guohua Lou; Liang Chen; Caixia Xia; Weina Wang; Jinjin Qi; Aichun Li; Liying Zhao; Zhi Chen; Min Zheng; Yanning Liu

    MiR-199a-3p (miR-199a) can enhance the chemosensitivity of hepatocellular carcinoma (HCC). Because of the easy degradation of miRNA by direct infusion, effective vehicle-mediated delivery of miR-199a may represent a new strategy for improving HCC chemotherapy. Considering mesenchymal stem cell (MSC)-derived exosomes as promising natural nanovectors for drug and molecule delivery, we aimed to determine whether exosomes from adipose tissue-derived MSCs (AMSCs) could be used to deliver miR-199a and improve HCC chemosensitivity. MiR-199a-modified AMSCs (AMSC-199a) were constructed by miR-199a lentivirus infection and puromycin selection. MiR-199-modified exosomes (AMSC-Exo-199a) were isolated from the supernatant of AMSC-199a and were assessed by transmission electron microscopy, nanoparticle tracking analysis, and flow cytometry analysis. The expression levels of miR-199a in HCC samples, AMSCs, exosomes, and HCC cells were quantified by real-time PCR. The effects of AMSC-Exo-199a on HCC chemosensitivity were determined by cell proliferation and apoptosis assays and by i.v. injection into orthotopic HCC mouse models with doxorubicin treatment. MTOR, p-4EBP1 and p-70S6K levels in HCC cells and tissues were quantified by Western blot. AMSC-Exo-199a had the classic characteristics of exosomes and could effectively mediate miR-199a delivery to HCC cells. Additionally, AMSC-Exo-199a significantly sensitized HCC cells to doxorubicin by targeting mTOR and subsequently inhibiting the mTOR pathway. Moreover, i.v.-injected AMSC-Exo-199a could distribute to tumor tissue and markedly increased the effect of Dox against HCC in vivo. AMSC-Exo-199a can be an effective vehicle for miR-199a delivery, and they effectively sensitized HCC to chemotherapeutic agents by targeting mTOR pathway. AMSC-Exo-199a administration may provide a new strategy for improving HCC chemosensitivity.

    更新日期:2020-01-02
  • Approaching the challenges of MKK3/p38delta MAPK targeting for therapeutic purpose in colorectal cancer
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-27
    Lorenzo Stramucci; Gianluca Bossi

    MKK3 is a member of the dual specificity kinase group specific upstream activator of p38 MAPK proteins. We originally identified MKK3 as mutant p53 (mutp53) gain-of-function (GOF) upregulated target gene in different tumor models. To deeply investigate the MKK3 functions in cancer, taking advantage of a panel of authenticated colorectal cancer (CRC) lines and primary colonocytes, we found that MKK3 activates specifically p38delta MAPK protein, which signaling is further triggered by 5-fluorouracil (5-FU) treatments, a largely adopted chemotherapeutic drug in CRC clinical practice. The overall achieved results proposed the MKK3/p38delta MAPK as relevant molecular axis involved in abrogating efficacy to 5-FU treatments in CRC. This commentary will provide an overall discussion of the results that have been achieved contextualizing them in the overview of the knowledge in the p38 MAPK field in cancer disease.

    更新日期:2019-12-30
  • Musashi2 contributes to the maintenance of CD44v6+ liver cancer stem cells via notch1 signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-30
    Xiju Wang; Ronghua Wang; Shuya Bai; Si Xiong; Yawen Li; Man Liu; Zhenxiong Zhao; Yun Wang; Yuchong Zhao; Wei Chen; Timothy R. Billiar; Bin Cheng

    Liver cancer stem cells (LCSCs) contribute to hepatocellular carcinoma (HCC) development, metastasis, and drug resistance. MSI2 and Notch1 signaling are involved in the maintenance of CSCs. However, it is unknown whether MSI2 and Notch1 are involved in the maintenance of CD44v6+ LCSCs. Therefore, we investigated the clinical significance and function of MSI2 and its relationship with Notch1 signaling in the maintenance of stemness properties in CD44v6+ LCSCs. The expression of MSI2 and CD44v6 were detected by fresh specimens and a HCC tissue microarray. The tissue microarray containing 82 HCC samples was used to analyze the correlation between CD44v6 and MSI2. CD44v6+/− cells were isolated using microbeads sorting. We explored the roles of MSI2 and Notch1 signaling in CD44v6+ LCSCs by sphere formation assay, transwell assay, clone formation assay in vitro, and xenograft tumor models in vivo. A Notch RT2 PCR Array, Co-immunoprecipitation, and RNA-immunoprecipitation were used to further investigate the molecular mechanism of MSI2 in activating Notch1 signaling. Here, we found MSI2 expression was positively correlated with high CD44v6 expression in HCC tissues, and further correlated with tumor differentiation. CD44v6+ cells isolated from HCC cell lines exhibited increased self-renewal, proliferation, migration and invasion, resistance to Sorafenib and tumorigenic capacity. Both MSI2 and Notch1 signaling were elevated in sorted CD44v6+ cells than CD44v6- cells and played essential roles in the maintenance of stemness of CD44v6+ LCSCs. Mechanically, MSI2 directly bound to Lunatic fringe (LFNG) mRNA and protein, resulting in Notch1 activation. Our results demonstrated that MSI2 maintained the stemness of CD44v6+ LCSCs by activating Notch1 signaling through the interaction with LFNG, which could be a potential molecular target for stem cell-targeted therapy for liver cancer.

    更新日期:2019-12-30
  • CircNFIX promotes progression of glioma through regulating miR-378e/RPN2 axis
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-30
    Chenyu Ding; Zanyi Wu; Honghai You; Hongliang Ge; Shufa Zheng; Yuanxiang Lin; Xiyue Wu; Zhangya Lin; Dezhi Kang

    Circular RNA nuclear factor I X (circNFIX) has been reported to play an important role in glioma progression. However, the mechanism by which circNFIX participates in glioma progression remains poorly understood. GERIA online were used to analyze the abnormally expressed genes in glioma tissues. The expression levels of circNFIX, microRNA (miR)-378e and Ribophorin-II (RPN2) were measured by quantitative real-time polymerase chain reaction or western blot. Cell cycle distribution, apoptosis, glycolysis, migration and invasion were determined by flow cytometry, special kit and trans-well assays, respectively. The target association between miR-378e and circNFIX or RPN2 was confirmed by luciferase reporter assay, RNA immunoprecipitation and pull-down. Xenograft model was established to investigate the role of circNFIX in vivo. The expression of circNFIX was enhanced in glioma tissues and cells compared with matched controls and high expression of circNFIX indicated poor outcomes of patients. Knockdown of circNFIX led to arrest of cell cycle, inhibition of glycolysis, migration and invasion and promotion of apoptosis in glioma cells. circNFIX was a sponge of miR-378e. miR-378e overexpression suppressed cell cycle process, glycolysis, migration and invasion but promoted apoptosis. miR-378e silence abated the suppressive role of circNFIX knockdown in glioma progression. RPN2 as a target of miR-378e was positively regulated via circNFIX by competitively sponging miR-378e. Silencing circNFIX decreased glioma xenograft tumor growth by regulating miR-378e/RPN2 axis. Knockdown of circNFIX inhibits progression of glioma in vitro and in vivo by increasing miR-378e and decreasing RPN2, providing a novel mechanism for understanding the pathogenesis of glioma.

    更新日期:2019-12-30
  • ROCK2 deprivation leads to the inhibition of tumor growth and metastatic potential in osteosarcoma cells through the modulation of YAP activity
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-26
    Cinzia Zucchini; Maria Cristina Manara; Camilla Cristalli; Marianna Carrabotta; Sara Greco; Rosa Simona Pinca; Cristina Ferrari; Lorena Landuzzi; Michela Pasello; Pier-Luigi Lollini; Marco Gambarotti; Davide Maria Donati; Katia Scotlandi

    The treatment of metastatic osteosarcoma (OS) remains a challenge for oncologists, and novel therapeutic strategies are urgently needed. An understanding of the pathways that regulate OS dissemination is required for the design of novel treatment approaches. We recently identified Rho-associated coiled-coil containing protein kinase 2 (ROCK2) as a crucial driver of OS cell migration. In this study, we explored the impact of ROCK2 disruption on the metastatic capabilities of OS cells and analyzed its functional relationship with Yes-associated protein-1 (YAP), the main transcriptional mediator of mechanotransduction signaling. The effects of ROCK2 depletion on metastasis were studied in NOD Scid gamma (NSG) mice injected with U-2OS cells in which ROCK2 expression had been stably silenced. Functional studies were performed in vitro in human U-2OS cells and in three novel cell lines derived from patient-derived xenografts (PDXs) by using standard methods to evaluate malignancy parameters and signaling transduction. The nuclear immunostaining of YAP and the evaluation of its downstream targets Cysteine Rich Angiogenic Inducer 6, Connective Tissue Growth Factor and Cyclin D1 by quantitative PCR were performed to analyze YAP activity. The effect of the expression and activity of ROCK2 and YAP on tumor progression was analyzed in 175 OS primary tumors. The silencing of ROCK2 markedly reduced tumor growth and completely abolished the metastatic ability of U-2OS cells. The depletion of ROCK2, either by pharmacological inhibition or silencing, induced a dose- and time-dependent reduction in the nuclear expression and transcriptional activity of YAP. The nuclear expression of YAP was observed in 80/175 (46%) tumor samples and was significantly correlated with worse patient prognosis and a higher likelihood of metastasis and death. The use of verteporfin, a molecule that specifically inhibits the TEAD–YAP association, remarkably impaired the growth and migration of OS cells in vitro. Moreover to inhibiting YAP activity, our findings indicate that verteporfin also affects the ROCK2 protein and its functions. We describe the functional connection between ROCK2 and YAP in the regulation of OS cell migration and metastasis formation. These data provide support for the use of verteporfin as a possible therapeutic option to prevent OS cell dissemination.

    更新日期:2019-12-27
  • HOXC-AS1-MYC regulatory loop contributes to the growth and metastasis in gastric cancer
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-23
    Yangyang Dong; Xinyu Li; Zhibin Lin; Wenbing Zou; Yan Liu; Huiyang Qian; Jing Jia

    Gastric cancer (GC) is one of the most prevalent and deadly malignancies worldwide. Accumulating reports have indicated the participation of long non-coding RNAs (lncRNAs) in the onset and progression of GC. GSE109476 data was utilized to screen out lncRNAs dysregulated in GC. Gene expressions were determined by qRT-PCR and western blot. Both in vitro and in vivo experiments were carried out to assess the function of HOXC-AS1 in GC. The association between genes was verified via RIP, ChIP, CoIP, RNA pull down and luciferase reporter assays, as appropriate. HOXC-AS1 was discovered to be upregulated in GC and located both in cytoplasm and in nucleus in GC cells. Functionally, inhibition of HOXC-AS1 restrained GC cell growth and metastasis both in vitro and in vivo. Moreover, HOXC-AS1 was proved to be trans-activated by c-MYC in GC. In return, HOXC-AS1 positively regulated MYC expression in GC through targeting miR-590-3p/MYC axis in cytoplasm and modulating BRG1/β-catenin complex-activated MYC transcription in nucleus. Furthermore, the rescue assays verified that MYC mediated HOXC-AS1-affected GC progression. Our research illustrated a feedback loop of HOXC-AS1-MYC in aggravating GC cell growth and metastasis, highlighting HOXC-AS1 as a promising target for GC diagnosis and treatment.

    更新日期:2019-12-23
  • Combination of the natural compound Periplocin and TRAIL induce esophageal squamous cell carcinoma apoptosis in vitro and in vivo: Implication in anticancer therapy
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-21
    Lujuan Han; Suli Dai; Zhirong Li; Cong Zhang; Sisi Wei; Ruinian Zhao; Hongtao Zhang; Lianmei Zhao; Baoen Shan

    Esophageal cancer is one of the most common malignant tumors in the world. With currently available therapies, only 20% ~ 30% patients can survive this disease for more than 5 years. TRAIL, a natural ligand for death receptors that can induce the apoptosis of cancer cells, has been explored as a therapeutic agent for cancers, but it has been reported that many cancer cells are resistant to TRAIL, limiting the potential clinical use of TRAIL as a cancer therapy. Meanwhile, Periplocin (CPP), a natural compound from dry root of Periploca sepium Bge, has been studied for its anti-cancer activity in a variety of cancers. It is not clear whether CPP and TRAIL can have activity on esophageal squamous cell carcinoma (ESCC) cells, or whether the combination of these two agents can have synergistic activity. We used MTS assay, flow cytometry and TUNEL assay to detect the effects of CPP alone or in combination with TRAIL on ESCC cells. The mechanism of CPP enhances the activity of TRAIL was analyzed by western blot, dual luciferase reporter gene assay and chromatin immunoprecipitation (ChIP) assay. The anti-tumor effects and the potential toxic side effects of CPP alone or in combination with TRAIL were also evaluated in vivo. In our studies, we found that CPP alone or in combination with TRAIL could inhibit the proliferation of ESCC cells and induce apoptosis, and we certificated that combination of two agents exert synergized functions. For the first time, we identified FoxP3 as a key transcriptional repressor for both DR4 and DR5. By down-regulating FoxP3, CPP increases the expression of DR4/DR5 and renders ESCC cells much more sensitive to TRAIL. We also showed that CPP reduced the expression of Survivin by inhibiting the activity of Wnt/β-catenin pathway. All these contributed to synergistic activity of CPP and TRAIL on ESCC cells in vitro and in vivo. Our data suggest that CPP and TRAIL could be further explored as potential therapeutic approach for esophageal cancer.

    更新日期:2019-12-21
  • MYCN and PRC1 cooperatively repress docosahexaenoic acid synthesis in neuroblastoma via ELOVL2
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-19
    Yi Ding; Jie Yang; Yawen Ma; Tengteng Yao; Xingyu Chen; Shengfang Ge; Lihua Wang; Xianqun Fan

    The MYCN amplification is a defining hallmark of high-risk neuroblastoma. Due to irregular oncogenes orchestration, tumor cells exhibit distinct fatty acid metabolic features from non-tumor cells. However, the function of MYCN in neuroblastoma fatty acid metabolism reprogramming remains unknown. Gas Chromatography-Mass Spectrometer (GC-MS) was used to find the potential target fatty acid metabolites of MYCN. Real-time PCR (RT-PCR) and clinical bioinformatics analysis was used to find the related target genes. The function of the identified target gene ELOVL2 on cell growth was detected through CCK-8 assay, Soft agar colony formation assay, flow Cytometry assay and mouse xenograft. Chromatin immunoprecipitation (ChIP) and Immunoprecipitation-Mass Spectrometer (IP-MS) further identified the target gene and the co-repressor of MYCN. The fatty acid profile of MYCN-depleted neuroblastoma cells identified docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid with anti-tumor activity, significantly increased after MYCN depletion. Compared with MYCN single-copy neuroblastoma cells, DHA level was significantly lower in MYCN-amplified neuroblastoma cells. RT-PCR and clinical bioinformatics analysis discovered that MYCN interfered DHA accumulation via ELOVL fatty acid elongase 2 (ELOVL2) which is a rate-limiting enzyme of cellular DHA synthesis. Enforced ELOVL2 expression in MYCN-amplified neuroblastoma cells led to decreased cell growth and counteracted the growth-promoting effect of MYCN overexpression both in vitro and vivo. ELOVL2 Knockdown showed the opposite effect in MYCN single-copy neuroblastoma cells. In primary neuroblastoma, high ELOVL2 transcription correlated with favorable clinical tumor biology and patient survival. The mechanism of MYCN-mediated ELOVL2 inhibition contributed to epigenetic regulation. MYCN recruited PRC1 (Polycomb repressive complex 1), catalysed H2AK119ub (histone 2A lysine 119 monoubiquitination) and inhibited subsequent ELOVL2 transcription. The tumor suppressive properties of DHA and ELOVL2 are repressed by the MYCN and PRC1 jointly, which suggests a new epigenetic mechanism of MYCN-mediated fatty acid regulation and indicates PRC1 inhibition as a potential novel strategy to activate ELOVL2 suppressive functions.

    更新日期:2019-12-20
  • Mortaparib, a novel dual inhibitor of mortalin and PARP1, is a potential drug candidate for ovarian and cervical cancers
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-19
    Jayarani F. Putri; Priyanshu Bhargava; Jaspreet Kaur Dhanjal; Tomoko Yaguchi; Durai Sundar; Sunil C. Kaul; Renu Wadhwa

    Mortalin is enriched in a large variety of cancers and has been shown to contribute to proliferation and migration of cancer cells in multiple ways. It has been shown to bind to p53 protein in cell cytoplasm and nucleus causing inactivation of its tumor suppressor activity in cancer cells. Several other activities of mortalin including mitochondrial biogenesis, ATP production, chaperoning, anti-apoptosis contribute to pro-proliferative and migration characteristics of cancer cells. Mortalin-compromised cancer cells have been shown to undergo apoptosis in in vitro and in vivo implying that it could be a potential target for cancer therapy. We implemented a screening of a chemical library for compounds with potential to abrogate cancer cell specific mortalin-p53 interactions, and identified a new compound (named it as Mortaparib) that caused nuclear enrichment of p53 and shift in mortalin from perinuclear (typical of cancer cells) to pancytoplasmic (typical of normal cells). Biochemical and molecular assays were used to demonstrate the effect of Mortaparib on mortalin, p53 and PARP1 activities. Molecular homology search revealed that Mortaparib is a novel compound that showed strong cytotoxicity to ovarian, cervical and breast cancer cells. Bioinformatics analysis revealed that although Mortaparib could interact with mortalin, its binding with p53 interaction site was not stable. Instead, it caused transcriptional repression of mortalin leading to activation of p53 and growth arrest/apoptosis of cancer cells. By extensive computational and experimental analyses, we demonstrate that Mortaparib is a dual inhibitor of mortalin and PARP1. It targets mortalin, PARP1 and mortalin-PARP1 interactions leading to inactivation of PARP1 that triggers growth arrest/apoptosis signaling. Consistent with the role of mortalin and PARP1 in cancer cell migration, metastasis and angiogenesis, Mortaparib-treated cells showed inhibition of these phenotypes. In vivo tumor suppression assays showed that Mortaparib is a potent tumor suppressor small molecule and awaits clinical trials. These findings report (i) the discovery of Mortaparib as a first dual inhibitor of mortalin and PARP1 (both frequently enriched in cancers), (ii) its molecular mechanism of action, and (iii) in vitro and in vivo tumor suppressor activity that emphasize its potential as an anticancer drug.

    更新日期:2019-12-20
  • Correction to: LncMAPK6 drives MAPK6 expression and liver TIC self-renewal
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-20
    Guanqun Huang; Hui Jiang; Yueming He; Ye Lin; Wuzheng Xia; Yuanwei Luo; Min Liang; Boyun Shi; Xinke Zhou; Zhixiang Jian

    In the original publication of this article [1], the author found an error in Fig. 2f. lncGPR107 should be changed to lncMAPK6, and the corrected Fig. 2 is shown below.

    更新日期:2019-12-20
  • Up-regulation of IGF2BP2 by multiple mechanisms in pancreatic cancer promotes cancer proliferation by activating the PI3K/Akt signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-18
    Xiaodong Xu; Yan Yu; Ke Zong; Pengwei Lv; Yuantin Gu

    The survival of pancreatic cancer patients remains poor. However, the underlying molecular mechanism and new therapeutic target of pancreatic cancer are still needed to be found. Many studies have shown that the IGF2 mRNA-binding protein 2 (IGF2BP2) plays oncogenic roles in cancers. However, the clinical significance, role and molecular mechanisms of IGF2BP2 in pancreatic cancer remain unclear. The expression of IGF2BP2 and miR-141 was detected in pancreatic cancer, and clinical significances were analyzed by statistical analysis. The function of IGF2BP2 and miR-141 was determined in vitro and in vivo, and the underlying mechanism was investigated. The gene copy number variation (CNV) of IGF2BP2 was analyzed based on The Cancer Genome Atlas (TCGA) dataset. microRNAs (miRNAs) regulating IGF2BP2 were predicted by online tools and confirmed by experiments. IGF2BP2 is overexpressed in pancreatic cancer tissues compared with control tissues. Upregulation of IGF2BP2 predicts shorter overall survival (OS) in pancreatic cancer patients by statistical analysis. IGF2BP2 overexpression is partially due to genomic amplification. Bioinformatics analyses and validation experiments showed that IGF2BP2 is a direct target of miR-141. A negative correlation between IGF2BP2 mRNA expression and the expression of miR-141 was observed in pancreatic cancer tissues and more importantly, reexpression of miR-141 rescued the oncogenic role of IGF2BP2. Moreover, upregulating IGF2BP2 expression promotes pancreatic cancer cell growth by activating the PI3K/Akt signaling pathway in vitro and in vivo. We comprehensively reveal the oncogenic role of IGF2BP2 in pancreatic cancer carcinogenesis and confirm that genomic amplification and the silencing of miR-141 contribute to its activation. Our findings highlight that IGF2BP2 may be a promising molecular target for the treatment of pancreatic cancer.

    更新日期:2019-12-19
  • Human bone marrow mesenchymal stem cells-derived microRNA-205-containing exosomes impede the progression of prostate cancer through suppression of RHPN2
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-17
    Shuangjian Jiang; Chengqiang Mo; Shengjie Guo; Jintao Zhuang; Bin Huang; Xiaopeng Mao

    Human bone marrow mesenchymal stem cells (hBMSCs) are implicated in cancer initiation and metastasis, sometimes by releasing exosomes that mediate cell communication by delivering microRNAs (miRNAs). This study aimed to investigate the physiological mechanisms by which exosomal miR-205 derived from hBMSCs may modulate the growth of prostate cancer cells. Microarray-based gene expression profiling of prostate cancer was adopted to identify differentially expressed genes and regulatory miRNAs, which identified the candidates RHPN2 and miR-205 as the study focus. Then the binding affinity between miR-205 and RHPN2 was identified using in silico analysis and luciferase activity detection. Prostate cancer cells were co-cultured with exosomes derived from hBMSCs treated with either miR-205 mimic or miR-205 inhibitor. Subsequently, prostate cancer cell proliferation, invasion, migration, and apoptosis were detected in vitro. The effects of hBMSCs-miR-205 on tumor growth were investigated in vivo. miR-205 was downregulated, while RHPN2 was upregulated in prostate cancer cells. RHPN2 was a target of miR-205, and upregulated miR-205 inhibited prostate cancer cell proliferation, invasion, and migration and promoted apoptosis by targeting RHPN2. Next, experiments demonstrated that hBMSCs-derived exosomes carrying miR-205 contributed to repressed prostate cancer cell proliferation, invasion, and migration and enhanced apoptosis. Furthermore, in vivo assays confirmed the inhibitory effects of hBMSCs-derived exosomal miR-205 on prostate cancer. The hBMSCs-derived exosomal miR-205 retards prostate cancer progression by inhibiting RHPN2, suggesting that miR-205 may present a predictor and potential therapeutic target for prostate cancer.

    更新日期:2019-12-18
  • Correction to: Role of mitochondria and cardiolipins in growth inhibition of breast cancer cells by retinoic acid
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-18
    Mineko Terao; Laura Goracci; Valentina Celestini; Mami Kurosaki; Marco Bolis; Alessandra Di Veroli; Arianna Vallerga; Maddalena Fratelli; Monica Lupi; Alessandro Corbelli; Fabio Fiordaliso; Maurizio Gianni; Gabriela Paroni; Adriana Zanetti; Gabriele Cruciani; Enrico Garattini

    In the original publication of this article [1], the images of Figs. 4 and 5 were exchanged and the legends of the two figures did not correspond due to a typesetting error.

    更新日期:2019-12-18
  • Triple blockade of EGFR, MEK and PD-L1 has antitumor activity in colorectal cancer models with constitutive activation of MAPK signaling and PD-L1 overexpression
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-16
    S. Napolitano; N. Matrone; A. L. Muddassir; G. Martini; A. Sorokin; V. De Falco; E. F. Giunta; D. Ciardiello; E. Martinelli; V. Belli; M. Furia; S. Kopetz; F. Morgillo; F. Ciardiello; T. Troiani

    Molecular mechanisms driving acquired resistance to anti-EGFR therapies in metastatic colorectal cancer (mCRC) are complex but generally involve the activation of the downstream RAS-RAF-MEK-MAPK pathway. Nevertheless, even if inhibition of EGFR and MEK could be a strategy for overcoming anti-EGFR resistance, its use is limited by the development of MEK inhibitor (MEKi) resistance. We have generated in vitro and in vivo different CRC models in order to underline the mechanisms of MEKi resistance. The three different in vitro MEKi resistant models, two generated by human CRC cells quadruple wild type for KRAS, NRAS, BRAF, PI3KCA genes (SW48-MR and LIM1215-MR) and one by human CRC cells harboring KRAS mutation (HCT116-MR) showed features related to the gene signature of colorectal cancer CMS4 with up-regulation of immune pathway as confirmed by microarray and western blot analysis. In particular, the MEKi phenotype was associated with the loss of epithelial features and acquisition of mesenchymal markers and morphology. The change in morphology was accompanied by up-regulation of PD-L1 expression and activation of EGFR and its downstream pathway, independently to RAS mutation status. To extend these in vitro findings, we have obtained mouse colon cancer MC38- and CT26-MEKi resistant syngeneic models (MC38-MR and CT26-MR). Combined treatment with MEKi, EGFR inhibitor (EGFRi) and PD-L1 inhibitor (PD-L1i) resulted in a marked inhibition of tumor growth in both models. These results suggest a strategy to potentially improve the efficacy of MEK inhibition by co-treatment with EGFR and PD-L1 inhibitors via modulation of host immune responses.

    更新日期:2019-12-17
  • Vitamin D, gut microbiota, and radiation-related resistance: a love-hate triangle
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-16
    Ruixue Huang; Jing Xiang; Pingkun Zhou

    Radiation resistance is a serious issue in radiotherapy. Increasing evidence indicates that the human gut microbiome plays a role in the development of radiation resistance. Vitamin D is an important supplement for cancer patients treated with radiotherapy. Against this background, this paper reviewed research regarding the associations among vitamin D, microbiota dysbiosis, and radiation resistance. A hypothesis is developed to describe the relationships among vitamin D, the gut microbiota, and radiotherapy outcomes. Radiotherapy changes the composition of the gut microbiota, which in turn influence the serum level of vitamin D, and its distribution and metabolism in the body. Alteration of vitamin D level influences the patient response to radiotherapy, where the underlying mechanisms may be associated with the intestinal microenvironment, immune molecules in the intestines, gut microbiome metabolites, and signaling pathways associated with vitamin D receptors. Our understanding of the contribution of vitamin D and the gut microbiota to radiotherapy outcomes has been increasing gradually. A better understanding of the relationships among vitamin D, the gut microbiota, and radiotherapy outcomes will shed more light on radiation resistance, and also promote the development of new strategies for overcoming it, thus addressing an important challenge associated with the currently available radiotherapy modalities for cancer patients.

    更新日期:2019-12-17
  • MicroRNA-338-3p suppresses ovarian cancer cells growth and metastasis: implication of Wnt/catenin beta and MEK/ERK signaling pathways
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-16
    Ruitao Zhang; Huirong Shi; Fang Ren; Wei Feng; Yuan Cao; Gailing Li; Zheying Liu; Pengcheng Ji; Minghui Zhang

    Downregulation of microRNA-338-3p (miR-338-3p) was detected in many malignant tumors, which indicated miR-338-3p might serve as a role of antioncogene in those cancers. The present study aimed to explore the roles of miR-338-3p in the growth and metastasis of ovarian cancer cells and elaborate the underlying possible molecular mechanism. Multiply biomedical databases query and KEGG pathway enrichment assay were used to infilter possible target genes and downstream pathways regulated by miR-338-3p. Overexpression miR-338-3p lentiviral vectors were transfected into ovarian cancer OVCAR-3 and OVCAR-8 cells, cell proliferation, migration and invasion were analyzed by MTT, colony formation, transwell, Matrigel assay and xenograft mouse model. One 3′-untranslated regions (UTRs) binding target gene of miR-338-3p, MACC1 (MET transcriptional regulator MACC1), and its regulated gene MET and downstream signaling pathway activities were examined by western blot. Biomedical databases query indicated that miR-338-3p could target MACC1 gene and regulate Met, downstream Wnt/Catenin beta and MEK/ERK pathways. Rescue of miR-338-3p could inhibit the proliferation, migration and invasion of ovarian cancer cells, and suppress the growth and metastasis of xenograft tumor. Restoration of miR-338-3p could attenuate MACC1 and Met overexpression induced growth, epithelial to mesenchymal transition (EMT) and activities of Wnt/Catenin beta and MEK/ERK signaling in vitro and in vivo. The present data indicated that restoration of miR-338-3p could suppress the growth and metastasis of ovarian cancer cells, which might due to the inhibition of proliferation and EMT induced by MACC1, Met and its downstream Wnt/Catenin beta and MEK/ERK signaling pathways.

    更新日期:2019-12-17
  • Ischemia reperfusion injury promotes recurrence of hepatocellular carcinoma in fatty liver via ALOX12-12HETE-GPR31 signaling axis
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-12
    Faji Yang; Yuheng Zhang; Haozhen Ren; Jinglin Wang; Longcheng Shang; Yang Liu; Wei Zhu; Xiaolei Shi

    Ischemia reperfusion injury (IRI) has been shown to increase the risk of tumor recurrence after liver surgery. Also, nonalcoholic fatty liver disease (NAFLD) is associated with increased HCC recurrence. ALOX12–12-HETE pathway is activated both in liver IRI and NASH. Also, ALOX12–12-HETE has been shown to mediate tumorigenesis and progression. Therefore, our study aims to investigate whether the ALOX12–12-HETE-GPR31 pathway involved in IRI induced HCC recurrence in NAFLD. HCC mouse model was used to mimic the HCC recurrence in NAFLD. Western Blot, qPCR, Elisa and Immunofluorescence analysis were conducted to evaluate the changes of multiple signaling pathways during HCC recurrence, including ALOX12–12-HETE axis, EMT, MMPs and PI3K/AKT/NF-κB signaling pathway. We also measured the expression and functional changes of GPR31 by siRNA. ALOX12–12-HETE pathway was activated in liver IRI and its activation was further enhanced in NAFLD, which induced more severe HCC recurrence in fatty livers than normal livers. Inhibition of ALOX12–12-HETE by ML355 reduced the HCC recurrence in fatty livers. In vitro studies showed that 12-HETE increased the expression of GPR31 and induced epithelial-mesenchymal transition (EMT) and matrix metalloprotein (MMPs) by activating PI3K/AKT/NF-κB pathway. Furthermore, knockdown of GPR31 in cancer cells inhibited the HCC recurrence in NAFLD. ALOX12–12-HETE-GPR31 played an important role in HCC recurrence and might be a potential therapeutic target to reduce HCC recurrence after surgery in fatty livers.

    更新日期:2019-12-13
  • N-myc downstream-regulated gene 1 inhibits the proliferation of colorectal cancer through emulative antagonizing NEDD4-mediated ubiquitylation of p21
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-12
    Sen Zhang; Chaoran Yu; Xiao Yang; Hiju Hong; Jiaoyang Lu; Wenjun Hu; Xiaohui Hao; Shuchun Li; Batuer Aikemu; Guang Yang; Zirui He; Luyang Zhang; Pei Xue; Zhenghao Cai; Junjun Ma; Lu Zang; Bo Feng; Fei Yuan; Jing Sun; Minhua Zheng

    N-myc downstream-regulated gene 1 (NDRG1) has been shown to play a key role in tumor metastasis. Recent studies demonstrate that NDRG1 can suppress tumor growth and is related to tumor proliferation; however, the mechanisms underlying these effects remain obscure. Immunohistochemistry (IHC) was used to detect NDRG1 and p21 protein expression in colorectal cancer tissue, and clinical significance of NDRG1 was also analyzed. CCK-8 assay, colony formation assay, flow cytometry, and xenograft model were used to assess the effect of NDRG1 on tumor proliferation in vivo and in vitro. The mechanisms underlying the effect of NDRG1 were investigated using western blotting, immunofluorescence, immunoprecipitation, and ubiquitylation assay. NDRG1 was down-regulated in CRC tissues and correlated with tumor size and patient survival. NDRG1 inhibited tumor proliferation through increasing p21 expression via suppressing p21 ubiquitylation. NDRG1 and p21 had a positive correlation both in vivo and in vitro. Mechanistically, E3 ligase NEDD4 could directly interact with and target p21 for degradation. Moreover, NDRG1 could emulatively antagonize NEDD4-mediated ubiquitylation of p21, increasing p21 expression and inhibit tumor proliferation. Our study could fulfill potential mechanisms of the NDRG1 during tumorigenesis and metastasis, which may serve as a tumor suppressor and potential target for new therapies in human colorectal cancer.

    更新日期:2019-12-13
  • Metformin in colorectal cancer: molecular mechanism, preclinical and clinical aspects
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-12
    Muhamad Noor Alfarizal Kamarudin; Md. Moklesur Rahman Sarker; Jin-Rong Zhou; Ishwar Parhar

    Growing evidence showed the increased prevalence of cancer incidents, particularly colorectal cancer, among type 2 diabetic mellitus patients. Antidiabetic medications such as, insulin, sulfonylureas, dipeptyl peptidase (DPP) 4 inhibitors and glucose-dependent insulinotropic peptide (GLP-1) analogues increased the additional risk of different cancers to diabetic patients. Conversely, metformin has drawn attention among physicians and researchers since its use as antidiabetic drug exhibited beneficial effect in the prevention and treatment of cancer in diabetic patients as well as an independent anticancer drug. This review aims to provide the comprehensive information on the use of metformin at preclinical and clinical stages among colorectal cancer patients. We highlight the efficacy of metformin as an anti-proliferative, chemopreventive, apoptosis inducing agent, adjuvant, and radio-chemosensitizer in various colorectal cancer models. This multifarious effects of metformin is largely attributed to its capability in modulating upstream and downstream molecular targets involved in apoptosis, autophagy, cell cycle, oxidative stress, inflammation, metabolic homeostasis, and epigenetic regulation. Moreover, the review highlights metformin intake and colorectal cancer risk based on different clinical and epidemiologic results from different gender and specific population background among diabetic and non-diabetic patients. The improved understanding of metformin as a potential chemotherapeutic drug or as neo-adjuvant will provide better information for it to be used globally as an affordable, well-tolerated, and effective anticancer agent for colorectal cancer.

    更新日期:2019-12-13
  • miR-624-5p promoted tumorigenesis and metastasis by suppressing hippo signaling through targeting PTPRB in osteosarcoma cells
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-11
    Yongjun Luo; Wei Liu; Pengyu Tang; Dongdong Jiang; Changjiang Gu; Yumin Huang; Fangyi Gong; Yuluo Rong; Dingfei Qian; Jian Chen; Zheng Zhou; Shujie Zhao; Jiaxing Wang; Tao Xu; Yongzhong Wei; Guoyong Yin; Jin Fan; Weihua Cai

    Accumulating evidence indicates that aberrant microRNA (miRNA) expression contributes to osteosarcoma progression. This study aimed to elucidate the association between miR-624-5p expression and osteosarcoma (OS) development and to investigate its underlying mechanism. We analyzed GSE65071 from the GEO database and found miR-624-5p was the most upregulated miRNA. The expression of miR-624-5p and its specific target gene were determined in human OS specimens and cell lines by RT-PCR and western blot. The effects of miR-624-5p depletion or ectopic expression on OS proliferation, migration and invasion were evaluated in vitro using CCK-8 proliferation assay, colony formation assay, transwell assay, would-healing assay and 3D spheroid BME cell invasion assay respectively. We investigated in vivo effects of miR-624-5p using a mouse tumorigenicity model. Besides, luciferase reporter assays were employed to identify interactions between miR-624-5p and its specific target gene. miR-624-5p expression was upregulated in OS cells and tissues, and overexpressing miR-624-5p led to a higher malignant level of OS, including cell proliferation, migration and invasion in vitro and in vivo. Protein tyrosine phosphatase receptor type B (PTPRB) was negatively correlated with miR-624-5p expression in OS tissues. Using the luciferase reporter assay and Western blotting, PTPRB was confirmed as a downstream target of miR-624-5p. PTPRB restored the effects of miR-624-5p on OS migration and invasion. The Hippo signaling pathway was identified as being involved in the miR-624-5p/PTPRB axis. In conclusion, our results suggest that miR-624-5p is a negative regulator of PTPRB and a risk factor for tumor metastasis in OS progression.

    更新日期:2019-12-11
  • Overexpressed ITGA2 promotes malignant tumor aggression by up-regulating PD-L1 expression through the activation of the STAT3 signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-09
    Dianyun Ren; Jingyuan Zhao; Yan Sun; Dan Li; Zibo Meng; Bo Wang; Ping Fan; Zhiqiang Liu; Xin Jin; Heshui Wu

    Recent studies have reported that Integrin alpha 2 (ITGA2) plays an essential role in tumor cell proliferation, invasion, metastasis, and angiogenesis. An abnormally expressed ITGA2 correlates with unfavorable prognoses in multiple types of cancer. However, the specific mechanism of how ITGA2 contributes to tumorigenesis remains unclear. The GEPIA web tool was used to find the clinical relevance of ITGA2 in cancer, and this significance was verified using Western blotting analysis of paired patient tissues and immunohistochemistry of the pancreatic cancer tissue. Functional assays, such as the MTS assay, colony formation assay, and transwell assay, were used to determine the biological role of ITGA2 in human cancer. The relationship between ITGA2 and programmed death-ligand 1 (PD-L1) was examined using Western blot analysis, RT-qPCR assay, and immunohistochemistry. The protein-protein interaction between ITGA2 and STAT3 was detected via co-immunoprecipitation. Our study showed that ITGA2 was markedly overexpressed in several malignant tumor cells and clinical tissues. Blocking ITGA2 inhibited the proliferation and invasion ability of cancer cells significantly, whereas overexpressed ITGA2 increased the degree of those processes considerably. Additionally, the RNA-seq assay indicated that ITGA2 transcriptionally regulated the expression of PD-L1 in pancreatic cancer. We also demonstrated that ITGA2 interacted with STAT3 and up-regulated the phosphorylation of STAT3; this interaction might involve the mechanism of ITGA2 inducing PD-L1 expression in cancer cells. Our results suggest that ITGA2 plays a critical role in cancer cell progression and the regulation of PD-L1 by activating the STAT3 pathway. We identified a novel mechanism by which ITGA2 plays a critical role in modulating cancer immune response by transcriptionally increasing the expression of PD-L1 in cancer cells. Thus, targeting ITGA2 is an effective method to enhance the efficacy of checkpoint immunotherapy against cancer.

    更新日期:2019-12-11
  • Long non-coding RNA PAXIP1-AS1 facilitates cell invasion and angiogenesis of glioma by recruiting transcription factor ETS1 to upregulate KIF14 expression
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-10
    Haiyang Xu; Guifang Zhao; Yu Zhang; Hong Jiang; Weiyao Wang; Donghai Zhao; Hongquan Yu; Ling Qi

    Gliomas are common life-threatening cancers, mainly due to their aggressive nature and frequent invasiveness and long non-coding RNAs (lncRNAs) are emerging as promising molecular targets. Therefore, we explored the regulatory mechanisms underlying the putative involvement of the lncRNA PAX-interacting protein 1- antisense RNA1/ETS proto-oncogene 1/kinesin family member 14 (PAXIP1-AS1/ETS1/KIF14) axis in glioma cell invasion and angiogenesis. Firstly, we identified differentially expressed lncRNA PAXIP1-AS1 as associated with glioma based on bioinformatic data. Then, validation experiments were conducted to confirm a high expression level of lncRNA PAXIP1-AS1 in glioma tissues and cells, accompanied by upregulated KIF14. We further examined the binding between lncRNA PAXIP1-AS1, KIF14 promoter activity, and transcription factor ETS1. Next, overexpression vectors and shRNAs were delivered to alter the expression of lncRNA PAXIP1-AS1, KIF14, and ETS1 to analyze their effects on glioma progression in vivo and in vitro. LncRNA PAXIP1-AS1 was mainly distributed in the nucleus of glioma cells. LncRNA PAXIP1-AS1 could upregulate the KIF14 promoter activity by recruiting transcription factor ETS1. Overexpression of lncRNA PAXIP1-AS1 enhanced migration, invasion, and angiogenesis of human umbilical vein endothelial cells in glioma by recruiting the transcription factor ETS1 to upregulate the expression of KIF14, which was further confirmed by accelerated tumor growth in nude mice. The key findings of this study highlighted the potential of the lncRNA PAXIP1-AS1/ETS1/KIF14 axis as a therapeutic target for glioma treatment, due to its role in controlling the migration and invasion of glioma cells and its angiogenesis.

    更新日期:2019-12-11
  • Arsenic sulfide induces RAG1-dependent DNA damage for cell killing by inhibiting NFATc3 in gastric cancer cells
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-10
    Ting Kang; Maolin Ge; Ruiheng Wang; Zhen Tan; Xiuli Zhang; Chuanying Zhu; Han Liu; Siyu Chen

    Arsenic sulfide was found to have potential anti-cancer activities, especially in gastric cancer. However, the underlying mechanism need to be further explored. This study was aimed to investigate the mechanism of arsenic compounds on gastric cancer. Gastric cancer cell lines were infected with lentiviral vector carrying shNFATc3 and/or treated with arsenic sulfide. MTT assay were performed to assess cell growth. Flow cytometer assays were used to detect cell cycle and reactive oxygen species (ROS) level of gastric cancer cells. Western blot was carried out to detect nuclear factor of activated T-cells, cytoplasmic 3 (NFATc3), cell cycle markers, DNA damage pathway protein expression as well as other protein expression in gastric cancer cell lines. The expression of recombination activating gene 1 (RAG1) in gastric cancer cell lines was determined by RNA-sequencing analyses and Real-Time qPCR. The effect of NFATc3 on RAG1 were determined by CHIP-qPCR assay. The effect of arsenic sulfide on AGS cells was evaluated in vivo. We show that arsenic sulfide as well as knockdown of NFATc3 resulted in increased double-strand DNA damage in gastric cancer cells by increasing the expression of RAG1, an endonuclease essential for immunoglobulin V(D) J recombination. Overexpression of NFATc3 blocked the expression of RAG1 expression and DNA damage induced by arsenic sulfide. Arsenic sulfide induced cellular oxidative stress to redistribute NFATc3, thereby inhibiting its transcriptional function, which can be reversed by N-acetyl-L-cysteine (NAC). We show that NFATc3 targets the promoter of RAG1 for transcriptional inhibition. We further showed that NFATc3 upregulation and RAG1 downregulation significantly associated with poor prognosis in patients with gastric cancer. Our in vivo experiments further confirmed that arsenic sulfide exerted cytotoxic activity against gastric cancer cells through inhibiting NFATc3 to activate RAG1 pathway. These results demonstrate that arsenic sulfide targets NFATc3 to induce double strand DNA break (DSB) for cell killing through activating RAG1 expression. Our results link arsenic compound to the regulation of DNA damage control and RAG1 expression as a mechanism for its cytotoxic effect.

    更新日期:2019-12-11
  • NOS1 inhibits the interferon response of cancer cells by S-nitrosylation of HDAC2
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-05
    Pengfei Xu; Shuangyan Ye; Keyi Li; Mengqiu Huang; Qianli Wang; Sisi Zeng; Xi Chen; Wenwen Gao; Jianping Chen; Qianbing Zhang; Zhuo Zhong; Ying Lin; Zhili Rong; Yang Xu; Bingtao Hao; Anghui Peng; Manzhao Ouyang; Qiuzhen Liu

    The dysfunction of type I interferon (IFN) signaling is an important mechanism of immune escape and metastasis in tumors. Increased NOS1 expression has been detected in melanoma, which correlated with dysfunctional IFN signaling and poor response to immunotherapy, but the specific mechanism has not been determined. In this study, we investigated the regulation of NOS1 on the interferon response and clarified the relevant molecular mechanisms. After stable transfection of A375 cells with NOS1 expression plasmids, the transcription and expression of IFNα-stimulated genes (ISGs) were assessed using pISRE luciferase reporter gene analysis, RT-PCR, and western blotting, respectively. The effect of NOS1 on lung metastasis was assessed in melanoma mouse models. A biotin-switch assay was performed to detect the S-nitrosylation of HDAC2 by NOS1. ChIP-qPCR was conducted to measure the binding of HDAC2, H4K16ac, H4K5ac, H3ac, and RNA polymerase II in the promoters of ISGs after IFNα stimulation. This effect was further evaluated by altering the expression level of HDAC2 or by transfecting the HDAC2-C262A/C274A site mutant plasmids into cells. The coimmunoprecipitation assay was performed to detect the interaction of HDAC2 with STAT1 and STAT2. Loss-of-function and gain-of-function approaches were used to examine the effect of HDAC2-C262A/C274A on lung metastasis. Tumor infiltrating lymphocytes were analyzed by flow cytometry. HDAC2 is recruited to the promoter of ISGs and deacetylates H4K16 for the optimal expression of ISGs in response to IFNα treatment. Overexpression of NOS1 in melanoma cells decreases IFNα-responsiveness and induces the S-nitrosylation of HDAC2-C262/C274. This modification decreases the binding of HDAC2 with STAT1, thereby reducing the recruitment of HDAC2 to the ISG promoter and the deacetylation of H4K16. Moreover, expression of a mutant form of HDAC2, which cannot be nitrosylated, reverses the inhibition of ISG expression by NOS1 in vitro and decreases NOS1-induced lung metastasis and inhibition of tumor infiltrating lymphocytes in a melanoma mouse model. This study provides evidence that NOS1 induces dysfunctional IFN signaling to promote lung metastasis in melanoma, highlighting NOS1-induced S-nitrosylation of HDAC2 in the regulation of IFN signaling via histone modification.

    更新日期:2019-12-06
  • Shisa3 brakes resistance to EGFR-TKIs in lung adenocarcinoma by suppressing cancer stem cell properties
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-04
    Jiahui Si; Yuanyuan Ma; Ji Wang Bi; Ying Xiong; Chao Lv; Shaolei Li; Nan Wu; Yue Yang

    Although EGFR tyrosine kinase inhibitors (EGFR-TKIs) are beneficial to lung adenocarcinoma patients with sensitive EGFR mutations, resistance to these inhibitors induces a cancer stem cell (CSC) phenotype. Here, we clarify the function and molecular mechanism of shisa3 as a suppressor that can reverse EGFR-TKI resistance and inhibit CSC properties. The suppresser genes involved in EGFR-TKI resistance were identified and validated by transcriptome sequencing, quantitative real-time PCR (qRT-PCR) and immunohistochemistry. Biological function analyses, cell half maximal inhibitory concentration (IC50), self-renewal, and migration and invasion capacities, were detected by CCK8, sphere formation and Transwell assays. Tumorigenesis and therapeutic effects were investigated in nonobese diabetic/severe combined immunodeficiency (nod-scid) mice. The underlying mechanisms were explored by Western blot and immunoprecipitation analyses. We found that low expression of shisa3 was related to EGFR-TKI resistance in lung adenocarcinoma patients. Ectopic overexpression of shisa3 inhibited CSC properties and the cell cycle in the lung adenocarcinoma cells resistant to gefitinib/osimertinib. In contrast, suppression of shisa3 promoted CSC phenotypes and the cell cycle in the cells sensitive to EGFR-TKIs. For TKI-resistant PC9/ER tumors in nod-scid mice, overexpressed shisa3 had a significant inhibitory effect. In addition, we verified that shisa3 inhibited EGFR-TKI resistance by interacting with FGFR1/3 to regulate AKT/mTOR signaling. Furthermore, combinational administration of inhibitors of FGFR/AKT/mTOR and cell cycle signaling could overcome EGFR-TKI resistance associated with shisa3-mediated CSC capacities in vivo. Taken together, shisa3 was identified as a brake to EGFR-TKI resistance and CSC characteristics, probably through the FGFR/AKT/mTOR and cell cycle pathways, indicating that shisa3 and concomitant inhibition of its regulated signaling may be a promising therapeutic strategy for reversing EGFR-TKI resistance.

    更新日期:2019-12-05
  • Discovery, characterization and potential roles of a novel NF-YAx splice variant in human neuroblastoma
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-05
    Lucia Cappabianca; Antonietta Rosella Farina; Lucia Di Marcotullio; Paola Infante; Daniele De Simone; Michela Sebastiano; Andrew Reay Mackay

    Identification of novel cancer-associated splice variants is of potential diagnostic, prognostic and therapeutic importance. NF-Y transcription factor is comprised of NF-YA, NF-YB and NF-YC subunits, binds inverted CCAAT-boxes in ≈70% of gene promoters, regulates > 1000 cancer-associated genes and proteins involved in proliferation, staminality, differentiation, apoptosis, metabolism and is subject to component alternative splicing. RT-PCR evaluation of alternative NF-YA splicing in primary human neuroblastomas (NBs), led to discovery of a novel NF-YAx splice variant, also expressed during mouse embryo development and induced by doxorubicin in NB cells. Here, we report the discovery and characterisation of NF-YAx and discus its potential roles in NB. NF-YAx cDNA was RT-PCR-cloned from a stage 3 NB (provided by the Italian Association of Haematology and Paediatric Oncology, Genova, IT), sequenced and expressed as a protein using standard methods and compared to known fully-spliced NF-YAl and exon B-skipped NF-YAs isoforms in: EMSAs for capacity to form NF-Y complexes; by co-transfection, co-immunoprecipitation and Western blotting for capacity to bind Sp1; by IF for localisation; in AO/EtBr cell-death and colony formation assays for relative cytotoxicity, and by siRNA knockdown, use of inhibitors and Western blotting for potential mechanisms of action. Stable SH-SY5Y transfectants of all three NF-YA isoforms were also propagated and compared by RT-PCR and Western blotting for differences in cell-death and stem cell (SC)-associated gene expression, in cell-death assays for sensitivity to doxorubicin and in in vitro proliferation, substrate-independent growth and in vivo tumour xenograft assays for differences in growth and tumourigenic capacity. NF-YAx was characterized as a novel variant with NF-YA exons B, D and partial F skipping, detected in 20% of NF-YA positive NBs, was the exclusive isoform in a stage 3 NB, expressed in mouse stage E11.5–14 embryos and induced by doxorubicin in SH-SY5Y NB cells. The NF-YAx protein exhibited nuclear localisation, competed with other isoforms in CCAAT box-binding NF-Y complexes but, in contrast to other isoforms, did not bind Sp1. NF-YAx expression in neural-related progenitor and NB cells repressed Bmi1 expression, induced KIF1Bβ expression and promoted KIF1Bβ-dependent necroptosis but in NB cells also selected tumourigenic, doxorubicin-resistant, CSC-like sub-populations, resistant to NF-YAx cytotoxicity. The discovery of NF-YAx in NBs, its expression in mouse embryos and induction by doxorubicin in NB cells, unveils a novel NF-YA splice mechanism and variant, regulated by and involved in development, genotoxic-stress and NB. NF-YAx substitution of other isoforms in NF-Y complexes and loss of capacity to bind Sp1, characterises this novel isoform as a functional modifier of NF-Y and its promotion of KIF1Bβ-dependent neural-lineage progenitor and NB cell necroptosis, association with doxorubicin-induced necroptosis and expression in mouse embryos coinciding with KIF1Bβ-dependent sympathetic neuroblast-culling, confirm a cytotoxic function and potential role in suppressing NB initiation. On the other hand, the in vitro selection of CSC-like NB subpopulations resistant to NF-YAx cytotoxicity not only helps to explain high-level exclusive NF-YAx expression in a stage 3 NB but also supports a role for NF-YAx in disease progression and identifies a potential doxorubicin-inducible mechanism for post-therapeutic relapse.

    更新日期:2019-12-05
  • PTPRD-inactivation-induced CXCL8 promotes angiogenesis and metastasis in gastric cancer and is inhibited by metformin
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-05
    Won Jung Bae; Ji Mi Ahn; Hye Eun Byeon; Seokwhi Kim; Dakeun Lee

    Protein tyrosine phosphatase receptor delta (PTPRD) is frequently inactivated in various types of cancers. Here, we explored the underlying mechanism of PTPRD-loss-induced cancer metastasis and investigated an efficient treatment option for PTPRD-inactivated gastric cancers (GCs). PTPRD expression was evaluated by immunohistochemistry. Microarray analysis was used to identify differentially expressed genes in PTPRD-inactivated cancer cells. Quantitative reverse transcription (qRT-PCR), western blotting, and/or enzyme-linked immunosorbent assays were used to investigate the PTPRD-CXCL8 axis and the expression of other related genes. An in vitro tube formation assay was performed using HUVECs. The efficacy of metformin was assessed by MTS assay. PTPRD was frequently downregulated in GCs and the loss of PTPRD expression was associated with advanced stage, worse overall survival, and a higher risk of distant metastasis. Microarray analysis revealed a significant increase in CXCL8 expression upon loss of PTPRD. This was validated in various GC cell lines using transient and stable PTPRD knockdown. PTPRD-loss-induced angiogenesis was mediated by CXCL8, and the increase in CXCL8 expression was mediated by both ERK and STAT3 signaling. Thus, specific inhibitors targeting ERK or STAT3 abrogated the corresponding signaling nodes and inhibited PTPRD-loss-induced angiogenesis. Additionally, metformin was found to efficiently inhibit PTPRD-loss-induced angiogenesis, decrease cell viability in PTPRD-inactivated cancers, and reverse the decrease in PTPRD expression. Thus, the PTPRD-CXCL8 axis may serve as a potential therapeutic target, particularly for the suppression of metastasis in PTPRD-inactivated GCs. Hence, we propose that the therapeutic efficacy of metformin in PTPRD-inactivated cancers should be further investigated.

    更新日期:2019-12-05
  • Correction to: Viral integration drives multifocal HCC during the occult HBV infection
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-12-04
    Xiao-Ping Chen; Xin Long; Wen-long Jia; Han-Jie Wu; Jing Zhao; Hui-Fang Liang; Arian Laurence; Jun Zhu; Dong Dong; Yan Chen; Long Lin; Yu-Dong Xia; Wei-Yang Li; Gui-Bo Li; Zhi-Kun Zhao; Kui Wu; Yong Hou; Jing-Jing Yu; Wei Xiao; Guo-Ping Wang; Peng-Cheng Zhu; Wei Chen; Ming-Zhou Bai; Yi-Xing Jian; Karsten Kristiansen; Qian Chen

    In the original publication of this article [1], Fig. 6 is wrong and the updated figure is shown below.

    更新日期:2019-12-04
  • Inhibition of lung cancer growth and metastasis by DHA and its metabolite, RvD1, through miR-138-5p/FOXC1 pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-29
    Xiaoming Bai; Jiaofang Shao; Sujin Zhou; Zhenggang Zhao; Fanghong Li; Rong Xiang; Allan Z. Zhao; Jinshun Pan

    Non small cell lung cancer (NSCLC) is one of the most common cancers in the world. DHA is known to be capable of suppressing NSCLC cell proliferation and metastasis. However, the mechanisms by which DHA exhibits its antitumor effects are unknown. Here we aimed to identify the effects and mechanisms of DHA and its metabolites on lung cancer cell growth and invasion. As measures of cell proliferation and invasion ability, the cell viability and transwell assays were used in vitro. Transgenic mfat-1 mice, which convert ω-6 PUFAs to ω-3 PUFAs, were used to detect the effect of endogenous DHA on tumor transplantation. An LC − MS/MS analysis identified the elevation of several eicosanoid metabolites of DHA. By using qPCR miRNA microarray, online prediction software, luciferase reporter assays and Western blot analysis, we further elucidated the mechanisms. Addition of exogenous DHA inhibited the growth and invasion in NSCLC cells in vitro. Endogenously produced DHA attenuated LLC-derived tumor growth and metastasis in the transgenic mfat-1 mice. Among the elevation of DHA metabolites, resolvin D1 (RvD1) significantly contributed to the inhibition in cell growth and invasion. MiRNA microarray revealed that the level of miR-138-5p was significantly increased after RvD1 treatment. MiR-138-5p mimics decreased cell viability and invasion; while miR-138-5p inhibitor abolished RvD1-mediated suppression of cell viability and invasion. The expression of FOXC1 was significantly reduced upon overexpression of miR-138-5p while luciferase reporter assay showed that FOXC1 was a direct target of miR-138-5p. In vivo, endogenous DHA by the mfat-1 transgene enhanced miR-138-5p expression and decreased FOXC1 expression. Furthermore, overexpression of FOXC1 reversed the inhibition in cell viability and invasion induced by RvD1 treatment. These data identified the RvD1/miR-138-5p/FOXC1 pathway as a novel mechanism by DHA and its metabolite, RvD1, and the potential of targeting such pathway as a therapeutic strategy in treating NSCLC.

    更新日期:2019-11-30
  • MFAP5 facilitates the aggressiveness of intrahepatic Cholangiocarcinoma by activating the Notch1 signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-27
    Jian-Hui Li; Xiao-Xu Zhu; Fu-Xi Li; Chen-Song Huang; Xi-Tai Huang; Jie-Qin Wang; Zhuo-Xing Gao; Shi-Jin Li; Qiong-Cong Xu; Wei Zhao; Xiao-Yu Yin

    Intrahepatic cholangiocarcinoma (ICC) is the second most common primary liver cancer. The dismal outcome of ICC patients is due to lack of early diagnosis, the aggressive biological behavior of ICC and the lack of effective therapeutic options. Early diagnosis and prognosis of ICC by non-invasive methods would be helpful in providing valuable information and developing effective treatment strategies. Expression of microfibrillar-associated protein 5 (MFAP5) in the serum of ICC patients was detected by ELISA. Human ICC specimens were immunostained by MFAP5 antibodies. The growth rate of human ICC cell lines treated with MFAP5 or MFAP5 shRNAs was examined by CCK8 and colony formation assays. Cell cycle analysis was performed with PI staining. The effect of MFAP5 inhibition was assessed by xenograft models in nude mice. RNA-seq and ATAC-seq analyses were used to dissect the molecular mechanism by which MFAP5 promoted ICC aggressiveness. We identified MFAP5 as a biomarker for the diagnosis and prognosis of ICC. Upregulated MFAP5 is a common feature in aggressive ICC patients’ tissues. Importantly, MFAP5 level in the serum of ICC patients and healthy individuals showed significant differential expression profiles. Furthermore, we showed that MFAP5 promoted ICC cell growth and G1 to S-phase transition. Using RNA-seq expression and ATAC-seq chromatin accessibility profiling of ICC cells with suppressed MFAP5 secretion, we showed that MFAP5 regulated the expression of genes involved in the Notch1 signaling pathway. Furthermore, FLI-06, a Notch signaling inhibitor, completely abolished the MFAP5-dependent transcriptional programs. Raised MFAP5 serum level is useful for differentiating ICC patients from healthy individuals, and could be helpful in ICC diagnosis, prognosis and therapies.

    更新日期:2019-11-28
  • Exosomal lncRNA ZFAS1 regulates esophageal squamous cell carcinoma cell proliferation, invasion, migration and apoptosis via microRNA-124/STAT3 axis
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-27
    Zhirong Li; Xuebo Qin; Wei Bian; Yishuai Li; Baoen Shan; Zhimeng Yao; Shujun Li

    In recent years, long non-coding RNAs (lncRNAs) are of great importance in development of different types of tumors, while the function of lncRNA ZFAS1 is rarely discussed in esophageal squamous cell carcinoma (ESCC). Therefore, we performed this study to explore the expression of exosomal lncRNA ZFAS1 and its molecular mechanism on ESCC progression. Expression of ZFAS1 and miR-124 in ESCC tissues was detected. LncRNA ZFAS1 was silenced to detect its function in the biological functions of ESCC cells. A stable donor and recipient culture model was established. Eca109 cells transfected with overexpressed and low expressed ZFAS1 plasmid and miR-124 inhibitor labeled by Cy3 were the donor cells, and then co-cultured with recipient cells to observe the transmission of Cy3-ZFAS1 between donor cells and recipient cells. The changes of cell proliferation, apoptosis, invasion, and migration in recipient cells were detected. The in vivo experiment was conducted for verifying the in vitro results. LncRNA ZFAS1 was upregulated and miR-124 was down-regulated in ESCC tissues. Silencing of ZFAS1 contributed to suppressed proliferation, migration, invasion and tumor growth in vitro and induced apoptosis of ESCC cells. LncRNA ZFAS1 was considered to be a competing endogenous RNA to regulate miR-124, thereby elevating STAT3 expression. Exosomes shuttled ZFAS1 stimulated proliferation, migration and invasion of ESCC cells and restricted their apoptosis with increased STAT3 and declined miR-124. Furthermore, in vivo experiment suggested that elevated ZFAS1-exo promoted tumor growth in nude mice. This study highlights that exosomal ZFAS1 promotes the proliferation, migration and invasion of ESCC cells and inhibits their apoptosis by upregulating STAT3 and downregulating miR-124, thereby resulting in the development of tumorigenesis of ESCC.

    更新日期:2019-11-28
  • RNA-binding protein KHSRP promotes tumor growth and metastasis in non-small cell lung cancer
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-27
    Mingxia Yan; Lei Sun; Jing Li; Huajian Yu; Hechun Lin; Tao Yu; Fangyu Zhao; Miaoxin Zhu; Lei Liu; Qin Geng; Hanwei Kong; Hongyu Pan; Ming Yao

    KH-type splicing regulatory protein (KHSRP) plays an important role in cancer invasion, but the relevant mechanism is not well known. In the present study, we investigated the function and potential molecular mechanism of KHSRP in non-small cell lung cancer (NSCLC) metastasis and elucidated its clinical significance. Isobaric tags for relative and absolute quantitation and the SWATH™ approach were combined with nanoliquid chromatography-tandem mass spectrometry analysis to identify metastasis-associated nucleoproteins in NSCLC. Real-time PCR and Western blot were used to screen for metastasis-associated candidate molecules. Gene knockdown and overexpression were used to investigate their functions and molecular mechanisms in lung cancer cells. Coimmunoprecipitation (Co-IP) experiments were performed to identify the interactions between candidate molecules and their interacting proteins. Gene expression and its association with multiple clinicopathologic characteristics were analyzed by immunohistochemistry (IHC) and Western blot in human lung cancer specimens. KHSRP was identified as a metastasis-associated candidate molecule. In NSCLC cell lines, knockdown of KHSRP significantly reduced lung cancer cell proliferation, migration, and invasion in vitro and in vivo, whereas overexpression of KHSRP did the opposite. Mechanistically, the protein heterogeneous nuclear ribonucleoprotein C (C1/C2) (HNRNPC) was identified to interact with KHSRP using Co-IP experiments. In NSCLC cell lines, overexpression of HNRNPC significantly promoted lung cancer cell proliferation, migration, and invasion in vitro and in vivo. KHSRP and HNRNPC may induce human lung cancer cell invasion and metastasis by activating the IFN-α-JAK-STAT1 signaling pathway. Drastically higher expression levels of KHSRP and HNRNPC were observed in lung cancer tissues compared to those in adjacent noncancerous tissues. Increased KHSRP and HNRNPC expression was significantly associated with advanced tumor stages and metastasis (both lymph node and distant). Kaplan-Meier survival analysis showed that patients with high KHSRP and HNRNPC expression levels were predicted to have the shortest survival times and to have a poor prognosis. KHSRP plays an important role in NSCLC metastasis and may serve as a potential prognostic marker and novel therapeutic target for lung cancer metastasis treatment.

    更新日期:2019-11-28
  • DNMT3b/OCT4 expression confers sorafenib resistance and poor prognosis of hepatocellular carcinoma through IL-6/STAT3 regulation
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-26
    Ssu-Chuan Lai; Yu-Ting Su; Ching-Chi Chi; Yung-Che Kuo; Kam-Fai Lee; Yu-Chih Wu; Pei-Chi Lan; Muh-Hwa Yang; Te-Sheng Chang; Yen-Hua Huang

    The inflammatory cytokine interleukin-6 (IL-6) is critical for the expression of octamer-binding transcription factor 4 (OCT4), which is highly associated with early tumor recurrence and poor prognosis of hepatocellular carcinomas (HCC). DNA methyltransferase (DNMT) family is closely linked with OCT4 expression and drug resistance. However, the underlying mechanism regarding the interplay between DNMTs and IL-6-induced OCT4 expression and the sorafenib resistance of HCC remains largely unclear. HCC tissue samples were used to examine the association between DNMTs/OCT4 expression levels and clinical prognosis. Serum levels of IL-6 were detected using ELISA assays (n = 144). Gain- and loss-of-function experiments were performed in cell lines and mouse xenograft models to determine the underlying mechanism in vitro and in vivo. We demonstrate that levels of DNA methyltransferase 3 beta (DNMT3b) are significantly correlated with the OCT4 levels in HCC tissues (n = 144), and the OCT4 expression levels are positively associated with the serum IL-6 levels. Higher levels of IL-6, DNMT3b, or OCT4 predicted early HCC recurrence and poor prognosis. We show that IL-6/STAT3 activation increases DNMT3b/1 and OCT4 in HCC. Activated phospho-STAT3 (STAT-Y640F) significantly increased DNMT3b/OCT4, while dominant negative phospho-STAT3 (STAT-Y705F) was suppressive. Inhibiting DNMT3b with RNA interference or nanaomycin A (a selective DNMT3b inhibitor) effectively suppressed the IL-6 or STAT-Y640F-induced increase of DNMT3b-OCT4 and ALDH activity in vitro and in vivo. The fact that OCT4 regulates the DNMT1 expressions were further demonstrated either by OCT4 forced expression or DNMT1 silence. Additionally, the DNMT3b silencing reduced the OCT4 expression in sorafenib-resistant Hep3B cells with or without IL-6 treatment. Notably, targeting DNMT3b with nanaomycin A significantly increased the cell sensitivity to sorafenib, with a synergistic combination index (CI) in sorafenib-resistant Hep3B cells. The DNMT3b plays a critical role in the IL-6-mediated OCT4 expression and the drug sensitivity of sorafenib-resistant HCC. The p-STAT3 activation increases the DNMT3b/OCT4 which confers the tumor early recurrence and poor prognosis of HCC patients. Findings from this study highlight the significance of IL-6-DNMT3b–mediated OCT4 expressions in future therapeutic target for patients expressing cancer stemness-related properties or sorafenib resistance in HCC.

    更新日期:2019-11-27
  • Forkhead box (FOX) G1 promotes hepatocellular carcinoma epithelial-Mesenchymal transition by activating Wnt signal through forming T-cell factor-4/Beta-catenin/FOXG1 complex
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-27
    Xingrong Zheng; Jiaxin Lin; Hewei Wu; Zhishuo Mo; Yunwen Lian; Peipei Wang; Zhaoxia Hu; Zhiliang Gao; Liang Peng; Chan Xie

    Forkhead box G1 (FOXG1) is a member of the Fox transcription factor family involved in regulation of many cancers. However, the role of FOXG1 in hepatocellular carcinogenesisis largely unclear. The present study aimed at examining the biological function and underlying mechanism of FOXG1 on hepatocellular carcinoma (HCC) tumor metastasis as well as its clinical significance. Levels of FOXG1 were determined by immunohistochemical and real-time PCR analysis in HCC cell lines and human HCC samples. The effect of FOXG1 on cancer cell invasion and metastasis was investigated in vitro and in vivo in either FOXG1-silenced or overexpressing human HCC cell lines. Immunoprecipitation and chromatin immunoprecipitation assays were performed to investigate the interaction of FOXG1, β-catenin, TCF4 and the effect on Wnt target-gene promoters. In human HCC, the level of FOXG1 progressively increased from surrounding non tumorous livers to HCC, reaching the highest levels in metastatic HCC. Furthermore, expression levels of FOXG1 directly correlated with cancer cell epithelial-mesenchymal transition (EMT) phenotype. In FOXG1-overexpressing cells, FOXG1 promotes the stabilization and nuclear accumulation of β-catenin by directly binding to β-catenin and it associates with the lymphoid enhancer factor/T cell factor proteins (LEF/TCFs) on Wnt responsive enhancers (WREs) in chromatin. The results show that FOXG1 plays a key role in mediating cancer cell metastasis through the Wnt/β-catenin pathway in HCC cells and predicts HCC prognosis after surgery. Targeting FOXG1 may provide a new approach for therapeutic treatment in the future.

    更新日期:2019-11-27
  • Pre-metastatic niche triggers SDF-1/CXCR4 axis and promotes organ colonisation by hepatocellular circulating tumour cells via downregulation of Prrx1
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-21
    Yujun Tang; Yishi Lu; Yuan Chen; Lei Luo; Lei Cai; Bangjian Peng; Wenbin Huang; Hangyu Liao; Liang Zhao; Mingxin Pan

    Circulating tumour cells (CTCs), especially mesenchymal CTCs, are important determinants of metastasis, which leads to most recurrence and mortality in hepatocellular carcinoma (HCC). However, little is known about the underlying mechanisms of CTC colonisation in pre-metastatic niches. Detection and classification of CTCs in patients were performed using the CanPatrol™ system. A lentiviral vector expressing Prrx1-targeting shRNA was constructed to generate a stable HCC cell line with low expression of Prrx1. The effect of Prrx1 knockdown on stemness, migration, and drug resistance of the cell line was assessed, including involvement of SDF-1/CXCR4 signalling. Promising clinical applications of an inhibitor of STAT3 tyrosine phosphorylation, C188–9, and specific blockade with CXCR4 antibody were explored. The number of mesenchymal CTCs in blood was closely associated with tumour recurrence or metastasis. Pre-metastatic niche-derived SDF-1 could downregulate Prrx1, which induced the stemness, drug resistance, and increased expression of CXCR4 in HCC cells through the STAT3 pathway in vitro. In vivo, mice bearing tumours of Prrx1 low-expressing cells had significantly shorter survival. In xenograft tumours and clinical samples, loss of Prrx1 was negatively correlated with increased expression of CXCR4 in lung metastatic sites compared with that in the primary foci. These findings demonstrate that decreased expression of Prrx1 stimulates SDF-1/CXCR4 signalling and contributes to organ colonisation with blood CTCs in HCC. STAT3 inhibition and specific blockade of CXCR4 have clinical potential as therapeutics for eliminating organ metastasis in advanced HCC.

    更新日期:2019-11-22
  • Long noncoding RNA 00976 promotes pancreatic cancer progression through OTUD7B by sponging miR-137 involving EGFR/MAPK pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-20
    Shan Lei; Zhiwei He; Tengxiang Chen; Xingjun Guo; Zhirui Zeng; Yiyi Shen; Jianxin Jiang

    Accumulation evidence indicates the vital role of long non-coding RNAs (lncRNAs) in tumorigenesis and the progression of malignant tumors, including pancreatic cancer (PC). However, the role and the molecular mechanism of long non-coding RNA 00976 is unclear in pancreatic cancer. In situ hybridization (ISH) and qRT-PCR was performed to investigate the association between linc00976 expression and the clinicopathological characteristics and prognosis of patients with PC. Subsequently, linc00976 over-expression vector and shRNAs were transfected into PC cells to up-regulate or down-regulate linc00976 expression. Loss- and gain-of function assays were performed to investigate the role of linc00976 in proliferation and metastasis in vitro and vivo. ITRAQ, bioinformatic analysis and rescue assay were used to illustrate the ceRNA mechanism network of linc00976/miR-137/OTUD7B and its downstream EGFR/MAPK signaling pathway. linc00976 expression was overexpressed in PC tissues and cell lines and was positively associated with poorer survival in patients with PC. Function studies revealed that linc00976 knockdown significantly suppressed cell proliferation, migration and invasion in vivo and in vitro, whereas its overexpression reversed these effects. Based on Itraq results and online database prediction, Ovarian tumor proteases OTUD7B was found as a downstream gene of linc00976, which deubiquitinated EGFR mediates MAPK signaling activation. Furthermore, Bioinformatics analysis and luciferase assays and rescue experiments revealed that linc00976/miR137/OTUD7B established the ceRNA network modulating PC cell proliferation and tumor growth. The present study demonstrates that linc00976 enhances the proliferation and invasion ability of PC cells by upregulating OTUD7B expression, which was a target of miR-137. Ultimately, OTUD7B mediates EGFR and MAPK signaling pathway, suggesting that linc00976/miR-137/OTUD7B/EGFR axis may act as a potential biomarker and therapeutic target for PC.

    更新日期:2019-11-21
  • Arf6-driven endocytic recycling of CD147 determines HCC malignant phenotypes
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-21
    Shanshan Qi; Linjia Su; Jing Li; Chuanshan Zhang; Zhe Ma; Guiqiu Liu; Qing Zhang; Guhe Jia; Yongjun Piao; Sihe Zhang

    Adhesion molecules distributed on the cell-surface depends upon their dynamic trafficking that plays an important role during cancer progression. ADP-ribosylation factor 6 (Arf6) is a master regulator of membrane trafficking. CD147, a tumor-related adhesive protein, can promote the invasion of liver cancer. However, the role of Arf6 in CD147 trafficking and its contribution to liver cancer progression remain unclear. Stable liver cancer cell lines with Arf6 silencing and over-expression were established. Confocal imaging, flow cytometry, biotinylation and endomembrane isolation were used to detect CD147 uptake and recycling. GST-pull down, gelatin zymography, immunofluorescence, cell adhesion, aggregation and tight junction formation, Transwell migration, and invasion assays were used to examine the cellular phenotypes. GEPIA bioinformatics, patient’s specimens and electronic records collection, and immunohistochemistry were performed to obtain the clinical relevance for Arf6-CD147 signaling. We found that the endocytic recycling of CD147 in liver cancer cells was controlled by Arf6 through concurrent Rab5 and Rab22 activation. Disruption of Arf6-mediated CD147 trafficking reduced the cell-matrix and cell-cell adhesion, weakened cell aggregation and junction stability, attenuated MMPs secretion and cytoskeleton reorganization, impaired HGF-stimulated Rac1 activation, and markedly decreased the migration and invasion of liver cancer cells. Moreover, high-expression of the Arf6-CD147 signaling components in HCC (hepatocellular carcinoma) was closely correlated with poor clinical outcome of patients. Our results revealed that Arf6-mediated CD147 endocytic recycling is required for the malignant phenotypes of liver cancer. The Arf6-driven signaling machinery provides excellent biomarkers or therapeutic targets for the prevention of liver cancer.

    更新日期:2019-11-21
  • Downregulation of class II phosphoinositide 3-kinase PI3K-C2β delays cell division and potentiates the effect of docetaxel on cancer cell growth
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-21
    Ouma Cisse; Muzthahid Quraishi; Federico Gulluni; Federica Guffanti; Ioanna Mavrommati; Methushaa Suthanthirakumaran; Lara C. R. Oh; Jessica N. Schlatter; Ambisha Sarvananthan; Massimo Broggini; Emilio Hirsch; Marco Falasca; Tania Maffucci

    Alteration of signalling pathways regulating cell cycle progression is a common feature of cancer cells. Several drugs targeting distinct phases of the cell cycle have been developed but the inability of many of them to discriminate between normal and cancer cells has strongly limited their clinical potential because of their reduced efficacy at the concentrations used to limit adverse side effects. Mechanisms of resistance have also been described, further affecting their efficacy. Identification of novel targets that can potentiate the effect of these drugs or overcome drug resistance can provide a useful strategy to exploit the anti-cancer properties of these agents to their fullest. The class II PI3K isoform PI3K-C2β was downregulated in prostate cancer PC3 cells and cervical cancer HeLa cells using selective siRNAs and the effect on cell growth was determined in the absence or presence of the microtubule-stabilizing agent/anti-cancer drug docetaxel. Mitosis progression was monitored by time-lapse microscopy. Clonogenic assays were performed to determine the ability of PC3 and HeLa cells to form colonies upon PI3K-C2β downregulation in the absence or presence of docetaxel. Cell multi-nucleation was assessed by immunofluorescence. Tumour growth in vivo was assessed using a xenograft model of PC3 cells upon PI3K-C2β downregulation and in combination with docetaxel. Downregulation of PI3K-C2β delays mitosis progression in PC3 and HeLa cells, resulting in reduced ability to form colonies in clonogenic assays in vitro. Compared to control cells, PC3 cells lacking PI3K-C2β form smaller and more compact colonies in vitro and they form tumours more slowly in vivo in the first weeks after cells implant. Stable and transient PI3K-C2β downregulation potentiates the effect of low concentrations of docetaxel on cancer cell growth. Combination of PI3K-C2β downregulation and docetaxel almost completely prevents colonies formation in clonogenic assays in vitro and strongly inhibits tumour growth in vivo. These data reveal a novel role for the class II PI3K PI3K-C2β during mitosis progression. Furthermore, data indicate that blockade of PI3K-C2β might represent a novel strategy to potentiate the effect of docetaxel on cancer cell growth.

    更新日期:2019-11-21
  • Upregulation of ERp57 promotes clear cell renal cell carcinoma progression by initiating a STAT3/ILF3 feedback loop
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-10-30
    Yan Liu; Jian-Xing Wang; Zi-Yuan Nie; Yue Wen; Xin-Ju Jia; Li-Na Zhang; Hui-Jun Duan; Yong-Hong Shi

    ERp57 dysfunction has been shown to contribute to tumorigenesis in multiple malignances. However, the role of ERp57 in clear cell renal carcinoma (ccRCC) remains unclear. Cell proliferation ability was measured by MTT and colony forming assays. Western blotting and quantitative real-time PCR (qRT-PCR) were performed to measure protein and mRNA expression. Co-immunoprecipitation (CoIP) and proximity ligation assay (PLA) were performed to detect protein-protein interaction. Chromatin immunoprecipitation (ChIP), ribonucleoprotein immunoprecipitation (RIP), and oligo pull-down were used to confirm DNA–protein and RNA–protein interactions. Promoter luciferase analysis was used to detect transcription factor activity. Here we found ERp57 was overexpressed in ccRCC tissues, and the higher levels of ERp57 were correlated with poor survival in patients with ccRCC. In vivo and in vitro experiments showed that ccRCC cell proliferation was enhanced by ERp57 overexpression and inhibited by ERp57 deletion. Importantly, we found ERp57 positively regulated ILF3 expression in ccRCC cells. Mechanically, ERp57 was shown to bind to STAT3 protein and enhance the STAT3-mediated transcriptional activity of ILF3. Furthermore, ILF3 levels were increased in ccRCC tissues and associated with poor prognosis. Interestingly, we revealed that ILF3 could bind to ERp57 and positively regulate its expression by enhancing its mRNA stability. Furthermore, ccRCC cell proliferation was moderated via the ERp57/STAT3/ILF3 feedback loop. In summary, our results indicate that the ERp57/STAT3/ILF3 feedback loop plays a key role in the oncogenesis of ccRCC and provides a potential therapeutic target for ccRCC treatment.

    更新日期:2019-11-20
  • SIRT4 silencing in tumor-associated macrophages promotes HCC development via PPARδ signalling-mediated alternative activation of macrophages
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-19
    Zhi Li; He Li; Zhi-Bo Zhao; Wei Zhu; Pan-Pan Feng; Xi-Wen Zhu; Jian-Ping Gong

    The activation of tumour-associated macrophages (TAMs) contributes to the progression of hepatocellular carcinoma (HCC). SIRT4 acts as a tumour suppressor of tumour growth by regulating cell metabolism, inflammation, and anti-tumourigenesis. However, the involvement of SIRT4 in the activation of TAMs is unknown. Based on previous findings, the expression of SIRT4 in distinct groups of TAMs as well as the effect of SIRT4 silencing on macrophage polarization was investigated. The expression of SIRT4 in HCC tissues and peritumour tissues was tested by qRT-PCR, western blotting and histological analysis. A Kaplan-Meier survival curve was generated based on the expression of SIRT4 in the HCC samples. Next, immunofluorescence staining was used to evaluate distinct groups of TAMs in human HCC samples, and the expression of SIRT4 in M1 and M2 TAMs was examined by flow cytometry. A homograft mouse model was used to assess the effect of SIRT4 silencing in TAMs on the development of HCC cells. SIRT4 was significantly downregulated in HCC tumour tissues, and the expression of SIRT4 in peritumour tissues was positively associated with survival in patients. We further found that downregulation of SIRT4 was associated with increased macrophage infiltration and a high ratio of M2/M1 macrophages in HCC peritumour tissues. Using gene interference, we found that SIRT4 silencing in TAMs significantly modulated the alternative activation of macrophages and promoted in vitro and in vivo HCC cell growth. Mechanistically, we revealed that HCM restricted the expression of SIRT4 in macrophages and promoted alternative activation of macrophages via the FAO-PPARδ-STAT3 axis. Furthermore, we also revealed that elevated MCP-1 expression induced by SIRT4 downregulation was responsible for increased TAM infiltration in peritumour tissues. Overall, our results demonstrate that downregulation of SIRT4 in TAMs modulates the alternative activation of macrophages and promotes HCC development via the FAO-PPARδ-STAT3 axis. These results could provide a new therapeutic target for the treatment of HCC.

    更新日期:2019-11-19
  • USP7 is a novel Deubiquitinase sustaining PLK1 protein stability and regulating chromosome alignment in mitosis
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-15
    Yuchong Peng; Youhong Liu; Yingxue Gao; Bowen Yuan; Xuli Qi; Yuxin Fu; Qianling Zhu; Tuoyu Cao; Songwei Zhang; Linglong Yin; Xiong Li

    The deubiquitinase USP7 has been identified as an oncogene with key roles in tumorigenesis and therapeutic resistance for a series of cancer types. Recently small molecular inhibitors have been developed to target USP7. However, the anticancer mechanism of USP7 inhibitors is still elusive. Cell viability or clonogenicity was tested by violet crystal assay. Cell apoptosis or cell cycle was analyzed by flow cytometry, and chromosome misalignment was observed by a fluorescent microscopy. The protein interaction of PLK1 and USP7 was detected by tandem affinity purification and high throughput proteomics, and further confirmed by co-immunoprecipitation, GST pull-down and protein co-localization. The correlation between USP7 level of tumor tissues and taxane-resistance was evaluated. Pharmacological USP7 inhibition by P5091 retarded cell proliferation and induced cell apoptosis. Further studies showed that P5091 induced cell cycle arrest at G2/M phase, and particularly induced chromosome misalignment, indicating the key roles of USP7 in mitosis. USP7 protein was detected in the PLK1-interacted protein complex. USP7 interacts with PLK1 protein through its PBD domain by catalytic activity. USP7 as a deubiquitinase sustained PLK1 protein stability via the C223 site, and inversely, USP7 inhibition by P5091 promoted the protein degradation of PLK1 through the ubiquitination-proteasome pathway. By overexpressing PLK1, USP7 that had been depleted by RNAi ceased to induce chromosome misalignment in mitosis and again supported cell proliferation and cell survival. Both USP7 and PLK1 were overexpressed in taxane-resistant cancer cells, and negatively correlated with the MP scores in tumor tissues. Either USP7 or PLK1 knockdown by RNAi significantly sensitized taxane-resistant cells to taxane cell killing. This is the first report that PLK1 is a novel substrate of USP7 deubiquitinase, and that USP7 sustained the protein stability of PLK1. USP7 inhibition induces cell apoptosis and cell cycle G2/M arrest, and overcomes taxane resistance by inducing the protein degradation of PLK1, resulting in chromosome misalignment in mitosis.

    更新日期:2019-11-15
  • CRISPR/Cas9-mediated knockout of NSD1 suppresses the hepatocellular carcinoma development via the NSD1/H3/Wnt10b signaling pathway
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-14
    Shuhua Zhang; Fan Zhang; Qing Chen; Chidan Wan; Jun Xiong; Jianqun Xu

    The NSD family of histone lysine methyltransferases have emerged as important biomarkers that participate in a variety of malignancies. Recent evidence has indicated that somatic dysregulation of the nuclear receptor binding SET domain-containing protein 1 (NSD1) is associated with the tumorigenesis in HCC, suggesting that NSD1 may serve as a prognostic target for this malignant tumor. However, its mechanism in human hepatocellular carcinoma (HCC), the major primary malignant tumor in the human liver, remains unclear. Hence, we investigated how NSD1 regulated HCC progression via regulation of the Wnt/β-catenin signaling pathway. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis was performed to identify the expression of NSD1 in HCC cells and clinically obtained tissues. The relationship between NSD1 expression and prognosis was analyzed by Kaplan-Meier survival curve. Further, a NSD1 knockout cell line was constructed by CRISPR/Cas9 genomic editing system, which was investigated in a battery of assays such as HCC cell proliferation, migration and invasion, followed by the investigation into NSD1 regulation on histone H3, Wnt10b and Wnt/β-catenin signaling pathway via ChIP. Finally, a nude mouse xenograft model was conducted in order to assess tumorigenesis affected by NSD1 knockout in vivo. NSD1 was overexpressed in HCC tissues and cell lines in association with poor prognosis. Knockout of NSD1 inhibited the proliferation, migration and invasion abilities of HCC cells. CRISPR/Cas9-mediated knockout of NSD1 promoted methylation of H3K27me3 and reduced methylation of H3K36me2, which inhibited Wnt10b expression. The results thereby indicated an inactivation of the Wnt/β-catenin signaling pathway suppressed cell proliferation, migration and invasion in HCC. Moreover, these in vitro findings were reproduced in vivo on tumor xenograft in nude mice. In conclusion, the study provides evidence that CRISPR/Cas9-mediated NSD1 knockout suppresses HCC cell proliferation and migration via the NSD1/H3/Wnt10b signaling pathway, suggesting that NSD1, H3 and Wnt10b may serve as potential targets for HCC.

    更新日期:2019-11-14
  • Programmed cell death ligand-1-mediated enhancement of hexokinase 2 expression is inversely related to T-cell effector gene expression in non-small-cell lung cancer
    J. Exp. Clin. Cancer Res. (IF 5.646) Pub Date : 2019-11-12
    Sehui Kim; Ji-Young Jang; Jaemoon Koh; Dohee Kwon; Young A. Kim; Jin Chul Paeng; Chan-Young Ock; Bhumsuk Keam; Miso Kim; Tae Min Kim; Dae Seog Heo; Doo Hyun Chung; Yoon Kyung Jeon

    We investigated the role of PD-L1 in the metabolic reprogramming of non-small cell lung cancer (NSCLC). Changes in glycolysis-related molecules and glycolytic activity were evaluated in PD-L1low and PD-L1high NSCLC cells after transfection or knockdown of PD-L1, respectively. Jurkat T-cell activation was assessed after co-culture with NSCLC cells. The association between PD-L1 and immune response-related molecules or glycolysis were analyzed in patients with NSCLC and The Cancer Genome Atlas (TCGA). Transfecting PD-L1 in PD-L1low cells enhanced hexokinase-2 (HK2) expression, lactate production, and extracellular acidification rates, but minimally altered GLUT1 and PKM2 expression and oxygen consumption rates. By contrast, knocking-down PD-L1 in PD-L1high cells decreased HK2 expression and glycolysis by suppressing PI3K/Akt and Erk pathways. Interferon-γ (IFNγ) secretion and activation marker expression was decreased in stimulated Jurkat T-cells when co-cultured with HK2-overexpressing vector-transfected tumor cells rather than empty vector-transfected tumor cells. Immunohistochemistry revealed that PD-L1 expression was positively correlated with HK2 expression in NSCLC (p < 0.001). In TCGA, HK2 exhibited a positive linear association with CD274 (PD-L1) expression (p < 0.001) but an inverse correlation with the expression of CD4, CD8A, and T-cell effector function-related genes in the CD274high rather than CD274low group. Consistently, there were fewer CD8+ T-cells in PD-L1positive/HK2high tumors compared to PD-L1positive/HK2low tumors in squamous cell carcinoma. PD-L1 enhances glycolysis in NSCLC by upregulating HK2, which might dampen anti-tumor immunity. PD-L1 may contribute to NSCLC oncogenesis by inducing metabolic reprogramming and immune checkpoint.

    更新日期:2019-11-13
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