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PDF file - 927K, The effect of Aurora kinases inhibitors on HL-60 and KG1a cell lines.

PDF file - 1715K, Aurora kinases inhibitors and mTOR inhibitors had synergistic effect on the expression of cleaved caspase 3 and p62.

PDF file - 1113K, Aurora kinases inhibitors had no effect with mTOR inhibitors on PBMCs.

PDF file - 1100K, Combination of Aurora kinases inhibitors and 2DG had minimal effect on PBMCs.

PDF file - 1470K, Aurora kinases inhibitors had synergistic effect with mTOR inhibitors on the glycolytic metabolism of U937 cells.

PDF file - 2735K, Aurora kinases inhibitors had synergistic effect with mTOR inhibitors on the proliferation of U937 and NB4 cells.

PDF file - 1081K, Cell viability detection in Aurora kinase A knockdown-U937 cells.

The mitotic kinase Aurora-A (Aur-A) is required to form the bipolar spindle and ensure accurate chromosome segregation before cell division. Aur-A dysregulation represents an oncogenic event that promotes tumor formation. Here, we report that Aur-A promotes breast cancer metastasis. Aur-A overexpression enhanced mammary cell migration by dephosphorylation and activation of cofilin, which facilitates actin reorganization and polymerization. Cofilin knockdown impaired Aur-A–driven cell migration and protrusion of the cell membrane. Conversely, overexpression of activated cofilin abrogated the effects of Aur-A knockdown on cell migration. Moreover, Aur-A overexpession increased the expression of the cofilin phosphatase Slingshot-1 (SSH1), contributing to cofilin activation and cell migration. We found that phosphatidylinositol 3-kinase (PI3K) inhibition blocked Aur-A–induced cofilin dephosphorylation, actin reorganization, and cell migration, suggesting crosstalk with PI3K signaling and a potential benefit of PI3K inhibition in tumors with deregulated Aur-A. Additionally, we found an association between Aur-A overexpression and cofilin activity in breast cancer tissues. Our findings indicate that activation of the cofilin-F-actin pathway contributes to tumor cell migration and metastasis enhanced by Aur-A, revealing a novel function for mitotic Aur-A kinase in tumor progression. Cancer Res; 70(22); 9118–28. ©2010 AACR.

Supplementary Figures 1-8, Tables 1-3, Methods from The Mitotic Kinase Aurora-A Induces Mammary Cell Migration and Breast Cancer Metastasis by Activating the Cofilin-F-actin Pathway

Small cell lung cancer (SCLC) is one of the most malignant types of lung cancer. Cancer stem cell (CSC) and tumor immune evasion are critical for the development of SCLC. We previously reported that NDR1 enhances breast CSC properties. NDR1 might also have a role in the regulation of immune responses. In the current study, we explore the function of NDR1 in the control of CSC properties and evasion of phagocytosis in SCLC. We find that NDR1 enhances the enrichment of the ALDEFLUOR

Lymphoma is one of the most curable types of cancer. However, drug resistance is the main challenge faced in lymphoma treatment. Peroxisomal acyl-CoA oxidase 1 (ACOX1) is the rate-limiting enzyme in fatty acid β-oxidation. Deregulation of ACOX1 has been linked to peroxisomal disorders and carcinogenesis in the liver. Currently, there is no information about the function of ACOX1 in lymphoma. In this study, we found that upregulation of ACOX1 promoted proliferation in lymphoma cells, while downregulation of ACOX1 inhibited proliferation and induced apoptosis. Additionally, overexpression of ACOX1 increased resistance to doxorubicin, while suppression of ACOX1 expression markedly potentiated doxorubicin-induced apoptosis. Interestingly, downregulation of ACOX1 promoted mitochondrial location of Bad, reduced mitochondrial membrane potential and provoked apoptosis by activating caspase-9 and caspase-3 related apoptotic pathway. Overexpression of ACOX1 alleviated doxorubicin-induced activation of caspase-9 and caspase-3 and decrease of mitochondrial membrane potential. Importantly, downregulation of ACOX1 increased p73, but not p53, expression. p73 expression was critical for apoptosis induction induced by ACOX1 downregulation. Also, overexpression of ACOX1 significantly reduced stability of p73 protein thereby reducing p73 expression. Thus, our study indicated that suppression of ACOX1 could be a novel and effective approach for treatment of lymphoma. [BMB Reports 2019; 52(9): 566-571].

Background Lymphoma is one of the most common hematologic malignancy. Drug resistance is the main obstacle faced in lymphoma treatment. Cancer stem cells are considered as the source of tumor recurrence, metastasis and drug resistance. The β-Asarone, a low-toxicity compound from the traditional medical herb Acorus calamus, has been shown to act as an anti-cancer reagent in various cancer types. However, the anti-cancer activities of β-Asarone in lymphoma have not been shown. Methods Cell counting assay was used to evaluate Raji cell proliferation. CCK8 assay was used to evaluate the cell viability. Annexin-V/PI staining and flow cytometry analysis were used to evaluate apoptosis. ALDEFLUOR assay was used to evaluate the stem-like population. Luciferase reporter assay was used to examine the activation of NF-κB signaling. Western blot and polymerase chain reaction (PCR) were used to determine the expression of interested genes. Results We showed that β-Asarone inhibited proliferation and induced apoptosis in Raji lymphoma cells in a dose-dependent manner. Additionally, β-Asarone functioned as a sensitizer of doxorubicin and resulted in synergistic effects on inhibition of proliferation and induction of apoptosis when combined with doxorubicin treatment. Interestingly, we found that β-Asarone also reduced the stem-like population of Raji lymphoma cells in a dose-dependent manner, and suppressed the expression of c-Myc and Bmi1. Importantly, β-Asarone abolished doxorubicin-induced enrichment of the stem-like population. In the mechanism study, we revealed that β-Asarone suppressed not only basal NF-κB activity but also Tumor necrosis factor α (TNF-α) induced NF-κB activity. Moreover, blocking NF-κB signaling inactivation was critical for β-Asarone induced apoptosis and inhibition of proliferation, but not for the effect on β-Asarone reduced stem-like population. In fact, β-Asarone suppressed stem-like population by destabilizing Bmi1 via a proteasome-mediated mechanism. Conclusions Our data suggested the application of β-Asarone to lower the toxic effect of doxorubicin and increase the sensitivity of doxorubicin in clinical treatment. More importantly, our data revealed a novel role of β-Asarone which could be used to eliminate stem-like population in lymphoma, implying that β-Asarone might reduce relapse and drug resistance. Electronic supplementary material The online version of this article (10.1186/s12935-019-0873-3) contains supplementary material, which is available to authorized users.

Additional file 1: Figure S1. β-Asarone inhibits NF-κB nuclear localization and the effect of the combination of β-Asarone and JSH-23 on apoptosis. A Raji cells were treated with 100 μM β-Asarone for 72 h. Cells were subjected to immunofluorescence staining. Representative images were shown. Images were magnified with a 100× objective. Scale bar, 10 μm. B Raji cells were treated with 400 μM β-Asarone and/or 7 μM JSH-23. Apoptosis was evaluated by the Annexin V-FITC/PI staining and flow cytometry analysis. Representative results are shown in the left panel and statistical results are shown in the right panel. Bar represents mean ± SD of three independent experiments (*p

// Min Yan 1, 2, * , Yuan-min Qian 1, * , Cai-feng Yue 1, 3 , Zi-feng Wang 1 , Bi-cheng Wang 1 , Wei Zhang 1 , Fei-meng Zheng 1, 4 , Quentin Liu 1 1 Sun Yat-sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Guangzhou, Institute of Cancer Stem Cell, Dalian, China 2 Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China 3 Department of Laboratory Medicine, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou, China 4 Department of Medical Oncology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, China * These authors have contributed equally to this work Correspondence to: Quentin Liu, email: liuq9@mail.sysu.edu.cn Keywords: histone deacetylases, p53, spindle, apoptosis, Nutlin-3 Received: February 25, 2016 Accepted: May 16, 2016 Published: June 8, 2016 ABSTRACT Histone deacetylases (HDACs) play crucial roles in the initiation and progression of cancer, offering a promising target for cancer therapy. HDACs inhibitor MGCD0103 (MGCD) exhibits effective anti-tumor activity by blocking proliferation and inducing cell death in malignant cells. However, the molecular mechanisms of HDACs inhibition induces cell death have not been well elucidated. In this study, we showed that MGCD effectively restored histone acetylation, suppressed cell growth and induced apoptosis in two-dimensional (2D) and three-dimensional (3D) cultured CNE1 and CNE2 nasopharyngeal carcinoma (NPC) cells. Importantly, MGCD arrested cell cycle at mitosis (M) phase with formation of multipolar spindles, which was associated with activated p53-mediated postmitotic checkpoint pathway to induce apoptotic cell death. Moreover, MGCD-induced apoptosis was decreased by inhibition of p53 using short interfering RNA (siRNA), suggesting that p53 was required for MGCD-induced cell apoptosis. Consistently, MGCD in combination with Nutlin-3, a MDM2 inhibitor showed synergistic effect on inducing apoptosis in 2D and 3D cultured CNE2 cells. Collectively, our data revealed that MGCD induced p53-dependent cell apoptosis following formation of multipolar spindles in NPC cells, suggesting the therapeutic potential of combinations of HDACs and MDM2 inhibitors for NPC treatment.

Background/Aims: Resistance of leukemia stem cells (LSCs) to chemotherapy in patients with acute myeloid leukemia (AML) causes relapse of disease. Hedgehog (Hh) signaling plays a critical role in the maintenance and differentiation of cancer stem cells. Yet its role in AML remains controversial. The purpose of the present study is to investigate the role of GLI1, the transcriptional activator of Hh signaling, in AML progenitor cells and to explore the anti-AML effects of GLI small-molecule inhibitor GANT61. Methods: The expression of GLI1 mRNA and protein were examined in AML progenitor cells and normal cells. The proliferation, colony formation, apoptosis and differentiation of AML progenitor cells were also analyzed in the presence of GANT61. Results: Kasumi-1 and KG1a cells, containing more CD34+ cells, expressed higher level of GLI1 compared to U937 and NB4 cells with fewer CD34+ cells. Consistently, a positive correlation between the protein levels of GLI1 and CD34 was validated in the bone marrow mononuclear cells (BMMC) of AML patients tested. GANT61 inhibited the proliferation and colony formation in AML cell lines. Importantly, GANT61 induced apoptosis in CD34+ enriched Kasumi-1 and KG1a cells, whereas it induced differentiation in U937 and NB4 cells. Furthermore, GANT61 enhanced the cytotoxicity of cytarabine (Ara-c) in primary CD34+ AML cells, indicating that inhibition of GLI1 could be a promising strategy to enhance chemosensitivity. Conclusions: The present findings suggested that Hh signaling was activated in AML progenitor cells. GLI1 acted as a potential target for AML therapy.

Drug resistance still represents a major obstacle to successful chronic myeloid leukemia (CML) treatment and novel compounds or strategies to override this challenging problem are urgently required. Here, we evaluated a novel compound AKI603 against oncogenic Aurora kinase A (Aur-A) in imatinib-resistant CML cells. We found that Aur-A was highly activated in imatinib-resistant KBM5-T315I cells. AKI603 significantly inhibited the phosphorylation of Aur-A kinase at Thr288, while had little inhibitory effect on BCR-ABL kinase in both KBM5 and KBM5-T315I cells. AKI603 inhibited cell viability, and induced cell cycle arrest with polyploidy accumulation in KBM5 and KBM5-T315I cells. Moreover, inhibition of Aur-A kinase by AKI603 suppressed colony formation capacity without promoting obvious apoptosis. Importantly, AKI603 promoted cell differentiation in both CML cell types. Thus, our study suggested the potential clinical use of small molecule Aurora kinase inhibitor AKI603 to overcome imatinib resistance in CML treatment.

Cancer stem-like cell (CS-like cell) is considered to be responsible for recurrence and drug resistance events in breast cancer, which makes it a potential target for novel cancer therapeutic strategy. The FDA approved flubendazole, has been widely used in the treatment of intestinal parasites. Here, we demonstrated a novel effect of flubendazole on breast CS-like cells. Flubendazole inhibited breast cancer cells proliferation in dose- and time-dependent manner and delayed tumor growth in xenograft models by intraperitoneal injection. Importantly, flubendazole reduced CD44high/CD24low subpopulation and suppressed the formation of mammosphere and the expression of self-renewal related genes including c-myc, oct4, sox2, nanog and cyclinD1. Moreover, we found that flubendazole induced cell differentiation and inhibited cell migration. Consistently, flubendazole reduced mesenchymal markers (β-catenin, N-cadherin and Vimentin) expression and induced epithelial and differentiation marker (Keratin 18) expression in breast cancer cells. Mechanism study revealed that flubendazole arrested cell cycle at G2/M phase and induced monopolar spindle formation through inhibiting tubulin polymerization. Furthermore, flubendazole enhanced cytotoxic activity of conventional therapeutic drugs fluorouracil and doxorubicin against breast cancer cells. In conclusion, our findings uncovered a remarkable effect of flubendazole on suppressing breast CS-like cells, indicating a novel utilization of flubendazole in breast cancer therapy.

Chemoresistance is a major cause of cancer treatment failure. Tumor-initiating cells (TIC) have attracted a considerable amount of attention due to their role in chemoresistance and tumor recurrence. Here, we evaluated the small-molecule Aurora kinase inhibitor AKI603 as a novel agent against TICs in breast cancer. AKI603 significantly inhibited Aurora-A (AurA) kinase and induced cell-cycle arrest. In addition, the intragastric administration of AKI603 reduced xenograft tumor growth. Interestingly, we found that breast cancer cells that were resistant to epirubicin expressed a high level of activated AurA and also have a high CD24Low/CD44High TIC population. The inhibition of AurA kinase by AKI603 abolished the epirubicin-induced enrichment of TICs. Moreover, AKI603 suppressed the capacity of cells to form mammosphere and also suppressed the expression of self-renewal genes (β-catenin, c-Myc, Sox2, and Oct4). Thus, our work suggests the potential clinical use of the small-molecule Aurora kinase inhibitor AKI603 to overcome drug resistance induced by conventional chemotherapeutics in breast cancer. Mol Cancer Ther; 13(8); 1991–2003. ©2014 AACR.

Tumor local invasion is the first step of metastasis cascade which remains the key obstacle for cancer therapy. Collective cell migration plays a critical role in tumor invading into surrounding tissues. In vitro assays fail to assess collective invasion in a real time manner. Herein we aim to develop a three-dimensional (3D) microfluidic cell invasion model to determine the dynamic process. In this model, collective invasion of breast cancer cells is induced by the concentration gradient of fetal bovine serum. We find that breast cancer cells adopt a collective movement rather than a random manner when the cells invade into extracellular matrix. The leading cells in the collective movement exhibit an increased expression of an Aurora kinase family protein - AURKA compared with the follower cells. Inhibition of AURKA kinase activity by VX680 or AKI603 significantly reduces the phosphorylation of ERK1/2 (Thr202/Tyr204) and collective cohort formation. Together, our study illustrates that AURKA acts as a potential therapeutic target for suppressing the process of tumor collective invasion. The 3D microfluidic cell invasion model is a reliable, measurable and dynamic platform for exploring potential drugs to inhibit tumor collective invasion.

Previously, we and others showed that mitotic Aurora-A kinase (Aur-A) was required for accurate mitotic entry and proper spindle assembly. In this study, we found that expression ofAur-Awas markedly elevated in bone marrow mononuclear cells (BMMCs) obtained from a significant portion of de novo acute myeloid leukemia (AML) patients. Targeting human primary AML cells with Aur-A kinase inhibitory VX-680 led to apoptotic cell death in a dose-dependent manner. Importantly, VX-680‐induced cell death was preferentially higher in Aur-A-high primary leukemic blasts compared with Aur-A-low AML (P < .001) or normal BMMCs (P < .001), suggesting the possible pharmacologic window in targeting Aurora kinase amongAur-A-high VX-680‐ sensitive leukemia patients. VX-680‐ induced cell death in AML cell lines was accompanied by formation of monopolar mitotic spindles, G2/M phase arrest, decreased phosphorylated(p)-Akt-1, and increased proteolytic cleavage of procaspase-3 and poly(ADP)ribose polymerase. Notably, VX-680 increased Bax/ Bcl-2 expression ratio, a favorable proapoptotic predictor for drug response and survival in AML. Lastly, VX-680 enhanced the cytotoxic effect of the chemotherapeutic agent etoposide (VP16) on AML cells. Together, we concluded that Aurora kinases were potentially therapeutic targets for AML and that Aur-A-high expression may serve as a differential marker for selective treatment. (Blood. 2008;111: 2854-2865)

// Dong-Ge Niu 1, 2, 3, * , Fei Peng 1, 2, * , Wei Zhang 1, 2 , Zhong Guan 4 , Hai-Dong Zhao 5 , Jing-Lin Li 1, 2, 3 , Kai-Li Wang 1, 2, 5 , Ting-Ting Li 1, 2 , Yan Zhang 1, 2 , Fei-Meng Zheng 1, 2 , Fan Xu 1, 2, 6 , Qian-Ni Han 1, 2, 3 , Peng Gao 3 , Qing-Ping Wen 3 , Quentin Liu 1, 2 1 Institute of Cancer Stem Cell, Dalian Medical University, Dalian 116044, China 2 Sun Yat-sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center of Cancer Medicine, Guangzhou 510060, China 3 Department of Anesthesia, The First Affiliated Hospital, Dalian Medical University, Dalian 116044, China 4 Department of Otorhinolaryngology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China 5 Department of Breast Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian 116023, China 6 Department of Clinical Pharmacology, College of Pharmacy, Dalian Medical University, Dalian 116044, China * These authors have contributed equally to this work Correspondence to: Quentin Liu, e-mail: liuq9@mail.sysu.edu.cn Qing-Ping Wen, e-mail: wqp.89@163.com Keywords: Morphine, cancer stem cell, chemoresistance, nalmefene, epithelial to mesenchymal transition Received: September 20, 2014 Accepted: December 11, 2014 Published: February 20, 2015 ABSTRACT Morphine is an opioid analgesic drug commonly used for pain relief in cancer patients. Here, we report that morphine enhances the mammosphere forming capacity and increases the expression of stemness-related transcription factors Oct4, Sox2 and Nanog. Treatment with morphine leads to enrichment of a side population fraction in MCF-7 cells and the CD44 + /CD24 −/low population in BT549 cells. Consistently, morphine activates Wnt/β-catenin signaling to induce epithelial to mesenchymal transition and promotes metastasis. Moreover, morphine decreases the sensitivity of traditional anti-cancer drugs in breast cancer cells. Nalmefene, an antagonist of morphine, reverses morphine-induced cancer stem cell properties and chemoresistance in breast cancer. In addition, nalmefene abolishes morphine enhancing tumorigenesis in a NOD/SCID mouse model. In conclusion, our findings demonstrate that morphine contributes to chemoresistance via expanding the population of cancer stem cells and promotes tumor growth, thereby revealing a novel role of morphine and providing some new guides in clinical use of morphine.

Background/Aim: P21, a multifunctional cell cycle-regulatory molecule, regulates apoptotic cell death. In this study we examined the effect of altered p21 expression on the sensitivity of acute myeloid leukemia cells in response to HDAC inhibitor SAHA treatment and investigated the underlying mechanism. Methods: Stably transfected HL60 cell lines were established in RPMI-1640 with supplementation of G-418. Cell viability was measured by MTT assay. Western blot was applied to assess the protein expression levels of target genes. Cell apoptosis was monitored by AnnexinV-PE/7AAD assay. Results: We showed HL60 cells that that didn't up-regulate p21 expression were more sensitive to SAHA-mediated apoptosis than NB4 and U937 cells that had increased p21 level. Enforced expression of p21 in HL60 cells reduced sensitivity to SAHA and blocked TRAIL-mediated apoptosis. Conversely, p21 silencing in NB4 cells enhanced SAHA-mediated apoptosis and lethality. Finally, we found that combined treatment with SAHA and rapamycin down-regulated p21 and enhanced apoptosis in AML cells. Conclusion: We conclude that up-regulated p21 expression mediates resistance to SAHA via inhibition of TRAIL apoptotic pathway. P21 may serve as a candidate biomarker to predict responsiveness or resistance to SAHA-based therapy in AML patients. In addition, rapamycin may be an effective agent to override p21-mediated resistance to SAHA in AML patients.

Lack of cellular differentiation is a key feature of nasopharyngeal carcinoma (NPC), but it also presents as a unique opportunity for intervention by differentiation therapy. Here using RNA-seq profiling analysis and functional assays, we demonstrate that reduced IKKα expression is responsible for the undifferentiated phenotype of NPC. Conversely, overexpression of IKKα induces differentiation and reduces tumorigenicity of NPC cells without activating NF-κB signalling. Importantly, we describe a mechanism whereby EZH2 directs IKKα transcriptional repression via H3K27 histone methylation on the IKKα promoter. The differentiation agent, retinoic acid, increases IKKα expression by suppressing EZH2-mediated H3K27 histone methylation, resulting in enhanced differentiation of NPC cells. In agreement, an inverse correlation between IKKα (low) and EZH2 (high) expression is associated with a lack of differentiation in NPC patient samples. Collectively, these findings demonstrate a role for IKKα in NPC differentiation and reveal an epigenetic mechanism for IKKα regulation, unveiling a new avenue for differentiation therapy.

Aurora kinases are overexpressed in large numbers of tumors and considered as potential therapeutic targets. In this study, we found that the Aurora kinases inhibitors MK-0457 (MK) and ZM447439 (ZM) induced polyploidization in acute myeloid leukemia (AML) cell lines. The level of glycolytic metabolism was significantly increased in the polyploidy cells, which were sensitive to glycolysis inhibitor 2-deoxy-D-glucose (2DG), suggesting that polyploidy cells might be eliminated by metabolism deprivation. Indeed, inhibition of mTOR pathway by mTOR inhibitors (rapamycin and PP242) or 2DG promoted not only apoptosis but also autophagy in the polyploidy cells induced by Aurora inhibitors. Mechanically, PP242 or2DGdecreased the level of glucose uptake and lactate production in polyploidy cells as well as the expression of p62/SQSTM1. Moreover, knockdown of p62/SQSTM1 sensitized cells to the Aurora inhibitor whereas overexpression of p62/SQSTM1 reduced drug efficacy. Thus, our results revealed that inhibition of mTOR pathway decreased the glycolytic metabolism of the polyploidy cells, and increased the efficacy of Aurora kinases inhibitors, providing a novel approach of combination treatment in AML. Mol Cancer Res; 11(11); 1326–36. ©2013 AACR.

The mitotic kinase Aurora-A (Aur-A) is required to form the bipolar spindle and ensure accurate chromosome segregation before cell division. Aur-A dysregulation represents an oncogenic event that promotes tumor formation. Here, we report that Aur-A promotes breast cancer metastasis. Aur-A overexpression enhanced mammary cell migration by dephosphorylation and activation of cofilin, which facilitates actin reorganization and polymerization. Cofilin knockdown impaired Aur-A–driven cell migration and protrusion of the cell membrane. Conversely, overexpression of activated cofilin abrogated the effects of Aur-A knockdown on cell migration. Moreover, Aur-A overexpession increased the expression of the cofilin phosphatase Slingshot-1 (SSH1), contributing to cofilin activation and cell migration. We found that phosphatidylinositol 3-kinase (PI3K) inhibition blocked Aur-A–induced cofilin dephosphorylation, actin reorganization, and cell migration, suggesting crosstalk with PI3K signaling and a potential benefit of PI3K inhibition in tumors with deregulated Aur-A. Additionally, we found an association between Aur-A overexpression and cofilin activity in breast cancer tissues. Our findings indicate that activation of the cofilin-F-actin pathway contributes to tumor cell migration and metastasis enhanced by Aur-A, revealing a novel function for mitotic Aur-A kinase in tumor progression. Cancer Res; 70(22); 9118–28. ©2010 AACR.

Elevated activity of the eukaryotic translation initiation factor 4E (eIF4E) plays crucial roles in tumorigenesis and disease progression by disproportionately increasing translation of mRNAs coding proteins that play significant roles in all aspects of malignancy, providing that eIF4E as an attractive target for therapeutic intervention. In this study, we showed that inhibition of eIF4E by small interfering RNAs (siRNA) resulted in cell cycle arrest and suppression of colony formation in MDA-MB-231 triple-negative (TN) breast cancer cells. Migration transwell assay revealed that repression of eIF4E effectively inhibited motility of MDA-MB-231 cancer cells. Importantly, we showed that silencing of eIF4E sensitized MDA-MB-231 cells to chemotherapeutic drugs of cisplatin, adriamycin, paclitaxel and docetaxel as assessed by MTT assay. Moreover, Western blot assay showed that eIF4E siRNA increased Bax/Bcl-2 ratio in MDA-MB-231 cells. Taken together, we showed that knockdown of eIF4E suppressed cell growth and migration, enhanced chemosensitivity, suggesting a potential therapeutic target in TN breast carcinoma.

Background There is increasing evidence that cancers contain their own stem-like cells, and particular attention has been paid to one subset of cancer-stem cells termed side population (SP). Stem cells under normal physical conditions are tightly controlled by their microenvironment, however, the regulatory role of the microenvironment surrounding cancer stem cells is not well characterized yet. In this study we found that the phenotype of SP can be "generated" by macrophage-like cells under conditioned culture. Furthermore the gene regulation pathway involved in cellular reprogramming process was investigated. Methods The selection and identification of SP in 50 CNE-2 single cell clones were performed by flow cytometry. The transwell assay and immunofluorescence staining were used to measure migration and cancer stem cell characters of non-SP single clone cells cultured with conditioned medium respectively. The subtraction suppression hybridization (SSH) technique and northern blotting analysis was applied to explore the pluripotency-associated genes under a unique epigenetic sub-microenvironment. Results Among 50 clones, only one did not possess SP subpopulation while others did. The non-SP cells induced by macrophage-like cells showed more aggressive characters, which increased cell migration compared with the control cells and showed some fraction of SP phenotype. These cells expressed distinguished level of pluripotency-associated genes such as ADP-ribosylation factor-like 6 interacting protein (ARMER), poly (rC) binding protein 1 (PCBP1) and pyruvate dehydrogenase E1-β subunit (PDHB) when subjected to the environment. Conclusion To our knowledge, this is the first study to demonstrate that non-SP single-clone cells can be induced to generate a SP phenotype when they are cultured with conditioned medium of macrophage-like cells, which is associated with the reactivation of pluripotency-associated genes.

Background The mitotic Aurora-A kinase exerts crucial functions in maintaining mitotic fidelity. As a bona fide oncoprotein, Aurora-A aberrant overexpression leads to oncogenic transformation. Yet, the mechanisms by which Aurora-A enhances cancer cell survival remain to be elucidated. Results Here, we found that Aurora-A overexpression was closely correlated with clinic stage and lymph node metastasis in tongue carcinoma. Aurora-A inhibitory VX-680 suppressed proliferation, induced apoptosis and markedly reduced migration in cancer cells. We further showed that insulin-like growth factor-1, a PI3K physiological activator, reversed VX-680-decreased cell survival and motility. Conversely, wortmannin, a PI3K inhibitor, combined with VX-680 showed a synergistic effect on inducing apoptosis and suppressing migration. In addition, Aurora-A inhibition suppressed Akt activation, and VX-680-induced apoptosis was attenuated by Myr-Akt overexpression, revealing a cross-talk between Aurora-A and PI3K pathway interacting at Akt activation. Significantly, we showed that suppression of Aurora-A decreased phosphorylated Akt and was associated with increased IkappaBα expression. By contrast, Aurora-A overexpression upregulated Akt activity and downregulated IkappaBα, these changes were accompanied by nuclear translocation of nuclear factor-κB and increased expression of its target gene Bcl-xL. Lastly, Aurora-A overexpression induced IkappaBα reduction was abrogated by suppression of Akt either chemically or genetically. Conclusion Taken together, our data established that Aurora-A, via activating Akt, stimulated nuclear factor-κB signaling pathway to promote cancer cell survival, and promised a novel combined chemotherapy targeting both Aurora-A and PI3K in cancer treatment.

Mitotic Aurora kinases are essential for accurate chromosome segregation during cell division. Forced overexpression of Aurora kinase results in centrosome amplification and multipolar spindles, causing aneuploidy, a hallmark of cancer. ZM447439 (ZM), an Aurora selective ATP-competitive inhibitor, interferes with the spindle integrity checkpoint and chromosome segregation. Here, we showed that inhibition of Aurora kinase by ZM reduced histone H3 phosphorylation at Ser10 in Hep2 carcinoma cells. Multipolar spindles were induced in these ZM-treated G(2)/M-arrested cells with accumulation of 4N/8N DNA, similar to cells with genetically suppressed Aurora-B. Cells subsequently underwent apoptosis, as assessed by cleavage of critical apoptotic associated protein PARP. Hep2 cells formed a tumor-like cell mass in 3-dimensional matrix culture; inhibition of Aurora kinase by ZM either destructed the preformed cell mass or prevented its formation, by inducing apoptotic cell death as stained for cleaved caspase-3. Lastly, ZM inhibition of Aurora kinase was potently in association with decrease of Akt phosphorylation at Ser473 and its substrates GSK3alpha/beta phosphorylation at Ser21 and Ser9. Together, we demonstrated that Aurora kinase served as a potential molecular target of ZM for more selective therapeutic cancer treatment.

Background Acute myeloid leukemia (AML) is an immunophenotypically heterogenous malignant disease, in which CD34 positivity is associated with poor prognosis. CD34+ AML cells are 10-15-fold more resistant to daunorubicin (DNR) than CD34- AML cells. Curcumin is a major component of turmeric that has shown cytotoxic activity in multiple cancers; however, its anti-cancer activity has not been well studied in DNR-insensitive CD34+ AML cells. The aim of this study was to therefore to explore curcumin-induced cytotoxicity in DNR-insensitive CD34+ AML cell lines (KG1a, Kasumi-1), DNR-sensitive U937 AML cells, and primary CD34+ AML bone-marrow-derived cells. Methods Primary human CD34+ cells were isolated from peripheral blood mononuclear cells or bone marrow mononuclear cells using a CD34 MicroBead kit. The growth inhibitory effects of curcumin were evaluated by MTT and colony-formation assays. Cell cycle distribution was examined by propidium iodide (PI) assay. Apoptosis was analyzed by Wright-Giemsa, Hoechst 33342 and Annexin-V/PI staining assays. The change in mitochondrial membrane potential (MMP) was examined by JC-1 staining and flow cytometry. Expression of apoptosis-related proteins was determined by reverse transcription-polymerase chain reaction and Western blotting. Short interfering RNA (siRNA) against Bcl-2 was used in CD34+ KG1a and Kasumi-1 cells incubated with/without DNR. Results Curcumin inhibited proliferation and induced apoptosis and G1/S arrest in both DNR-insensitive KG1a, Kasumi-1 and DNR-sensitive U937 cells. Curcumin-induced apoptosis was associated with reduced expression of both Bcl-2 mRNA and protein, subsequent loss of MMP, and activation of caspase-3 followed by PARP degradation. Curcumin synergistically enhanced the cytotoxic effect of DNR in DNR-insensitive KG1a and Kasumi-1 cells, consistent with decreased Bcl-2 expression. Accordingly, siRNA against Bcl-2 increased the susceptibility of KG1a and Kasumi-1 cells to DNR-induced apoptosis. More importantly, curcumin suppressed Bcl-2 expression, selectively inhibited proliferation and synergistically enhanced the cytotoxicity of DNR in primary CD34+ AML cells, while showing limited lethality in normal CD34+ hematopoietic progenitors. Conclusion Curcumin down-regulates Bcl-2 and induces apoptosis in DNR-insensitive CD34+ AML cell lines and primary CD34+ AML cells.