Showing papers in "Oncogene in 2010"

EMT, cancer stem cells and drug resistance: an emerging axis of evil in the war on cancer

Abstract: Tumors are cellularly and molecularly heterogeneous, with subsets of undifferentiated cancer cells exhibiting stem cell-like features (CSCs). Epithelial to mesenchymal transitions (EMT) are transdifferentiation programs that are required for tissue morphogenesis during embryonic development. The EMT process can be regulated by a diverse array of cytokines and growth factors, such as transforming growth factor (TGF)-β, whose activities are dysregulated during malignant tumor progression. Thus, EMT induction in cancer cells results in the acquisition of invasive and metastatic properties. Recent reports indicate that the emergence of CSCs occurs in part as a result of EMT, for example, through cues from tumor stromal components. Recent evidence now indicates that EMT of tumor cells not only causes increased metastasis, but also contributes to drug resistance. In this review, we will provide potential mechanistic explanations for the association between EMT induction and the emergence of CSCs. We will also highlight recent studies implicating the function of TGF-β-regulated noncoding RNAs in driving EMT and promoting CSC self-renewal. Finally we will discuss how EMT and CSCs may contribute to drug resistance, as well as therapeutic strategies to overcome this clinically.

Defining the role of hypoxia-inducible factor 1 in cancer biology and therapeutics

Abstract: Adaptation of cancer cells to their microenvironment is an important driving force in the clonal selection that leads to invasive and metastatic disease. O2 concentrations are markedly reduced in many human cancers compared with normal tissue, and a major mechanism mediating adaptive responses to reduced O2 availability (hypoxia) is the regulation of transcription by hypoxia-inducible factor 1 (HIF-1). This review summarizes the current state of knowledge regarding the molecular mechanisms by which HIF-1 contributes to cancer progression, focusing on (1) clinical data associating increased HIF-1 levels with patient mortality; (2) preclinical data linking HIF-1 activity with tumor growth; (3) molecular data linking specific HIF-1 target gene products to critical aspects of cancer biology and (4) pharmacological data showing anticancer effects of HIF-1 inhibitors in mouse models of human cancer.

Q's next: the diverse functions of glutamine in metabolism, cell biology and cancer.

Abstract: Several decades of research have sought to characterize tumor cell metabolism in the hope that tumor-specific activities can be exploited to treat cancer. Having originated from Warburg's seminal observation of aerobic glycolysis in tumor cells, most of this attention has focused on glucose metabolism. However, since the 1950s cancer biologists have also recognized the importance of glutamine (Q) as a tumor nutrient. Glutamine contributes to essentially every core metabolic task of proliferating tumor cells: it participates in bioenergetics, supports cell defenses against oxidative stress and complements glucose metabolism in the production of macromolecules. The interest in glutamine metabolism has been heightened further by the recent findings that c-myc controls glutamine uptake and degradation, and that glutamine itself exerts influence over a number of signaling pathways that contribute to tumor growth. These observations are stimulating a renewed effort to understand the regulation of glutamine metabolism in tumors and to develop strategies to target glutamine metabolism in cancer. In this study we review the protean roles of glutamine in cancer, both in the direct support of tumor growth and in mediating some of the complex effects on whole-body metabolism that are characteristic of tumor progression.

Immune infiltration in human tumors: a prognostic factor that should not be ignored.

Abstract: The natural history of a tumor includes phases of 'in situ' growth, invasion, extravasation and metastasis. During these phases, tumor cells interact with their microenvironment and are influenced by signals coming from stromal, endothelial, inflammatory and immune cells. Indeed, tumors are often infiltrated by various numbers of lymphocytes, macrophages or mast cells. It is generally believed that the latter produce factors that maintain chronic inflammation and promote tumor growth, whereas lymphocytes may control cancer outcome, as evidenced in mouse models. In this study, we analyze data from large cohorts of human tumors, clearly establishing that infiltration of the primary tumor by memory T cells, particularly of the Th1 and cytotoxic types, is the strongest prognostic factor in terms of freedom from disease and overall survival at all stages of clinical disease. We review data suggesting that tertiary lymphoid structures adjacent to tumors and composed of mature dendritic cells (T and B cells organized as germinal centers) may be the site of an antitumor reaction. We propose an immune scoring based on the type, density and location of lymphocyte infiltrates as a novel prognostic factor for use in addition to tumor node metastasis staging to predict disease-free survival and to aid in decisions regarding adjuvant therapies in early stage human cancers.

Immunogenic death of colon cancer cells treated with oxaliplatin.

Abstract: Both the pre-apoptotic exposure of calreticulin (CRT) and the post-apoptotic release of high-mobility group box 1 protein (HMGB1) are required for immunogenic cell death elicited by anthracyclins. Here, we show that both oxaliplatin (OXP) and cisplatin (CDDP) were equally efficient in triggering HMGB1 release. However, OXP, but not CDDP, stimulates pre-apoptotic CRT exposure in a series of murine and human colon cancer cell lines. Subcutaneous injection of OXP-treated colorectal cancer (CRC), CT26, cells induced an anticancer immune response that was reduced by short interfering RNA-mediated depletion of CRT or HMGB1. In contrast, CDDP-treated CT26 cells failed to induce anticancer immunity, unless recombinant CRT protein was absorbed into the cells. CT26 tumors implanted in immunocompetent mice responded to OXP treatment in vivo, and this therapeutic response was lost when CRT exposure by CT26 cells was inhibited or when CT26 cells were implanted in immunodeficient mice. The knockout of toll-like receptor 4 (TLR4), the receptor for HMGB1, also resulted in a deficient immune response against OXP-treated CT26 cells. In patients with advanced (stage IV, Duke D) CRC, who received an OXP-based chemotherapeutic regimen, the loss-of-function allele of TLR4 (Asp299Gly in linkage disequilibrium with Thr399Ile, reducing its affinity for HMGB1) was as prevalent as in the general population. However, patients carrying the TLR4 loss-of-function allele exhibited reduced progression-free and overall survival, as compared with patients carrying the normal TLR4 allele. In conclusion, OXP induces immunogenic death of CRC cells, and this effect determines its therapeutic efficacy in CRC patients.

The role of signaling pathways in the development and treatment of hepatocellular carcinoma.

Abstract: Hepatocellular carcinoma (HCC) is a highly prevalent, treatment-resistant malignancy with a multifaceted molecular pathogenesis. Current evidence indicates that during hepatocarcinogenesis, two main pathogenic mechanisms prevail: (1) cirrhosis associated with hepatic regeneration after tissue damage caused by hepatitis infection, toxins (for example, alcohol or aflatoxin) or metabolic influences, and (2) mutations occurring in single or multiple oncogenes or tumor suppressor genes. Both mechanisms have been linked with alterations in several important cellular signaling pathways. These pathways are of interest from a therapeutic perspective, because targeting them may help to reverse, delay or prevent tumorigenesis. In this review, we explore some of the major pathways implicated in HCC. These include the RAF/MEK/ERK pathway, phosphatidylinositol-3 kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway, WNT/beta-catenin pathway, insulin-like growth factor pathway, hepatocyte growth factor/c-MET pathway and growth factor-regulated angiogenic signaling. We focus on the role of these pathways in hepatocarcinogenesis, how they are altered, and the consequences of these abnormalities. In addition, we also review the latest preclinical and clinical data on the rationally designed targeted agents that are now being directed against these pathways, with early evidence of success.

The Role of COX-2 in Intestinal Inflammation and Colorectal Cancer

Abstract: Colorectal cancer (CRC) is a heterogeneous disease, including at least three major forms: hereditary, sporadic and colitis-associated CRC. A large body of evidence indicates that genetic mutations, epigenetic changes, chronic inflammation, diet and lifestyle are the risk factors for CRC. As elevated cyclooxygenase-2 (COX-2) expression was found in most CRC tissue and is associated with worse survival among CRC patients, investigators have sought to evaluate the effects of nonsteroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors (COXIBs) on CRC. The epidemiological studies, clinical trials and animal experiments indicate that NSAIDs are among the most promising chemopreventive agents for this disease. NSAIDs exert their anti-inflammatory and antitumor effects primarily by reducing prostaglandin production by inhibition of COX-2 activity. In this review, we highlight breakthroughs in our understanding of the roles of COX-2 in CRC and inflammatory bowel disease. These recent data provide a rationale for re-evaluating COX-2 as both the prognostic and the predictive marker in a wide variety of malignancies and for renewing the interest in evaluating relative benefits and risk of COXIBs in appropriately selected patients for cancer prevention and treatment.

Extracellular adenosine triphosphate and adenosine in cancer.

Abstract: Adenosine triphosphate (ATP) is actively released in the extracellular environment in response to tissue damage and cellular stress. Through the activation of P2X and P2Y receptors, extracellular ATP enhances tissue repair, promotes the recruitment of immune phagocytes and dendritic cells, and acts as a co-activator of NLR family, pyrin domain-containing 3 (NLRP3) inflammasomes. The conversion of extracellular ATP to adenosine, in contrast, essentially through the enzymatic activity of the ecto-nucleotidases CD39 and CD73, acts as a negative-feedback mechanism to prevent excessive immune responses. Here we review the effects of extracellular ATP and adenosine on tumorigenesis. First, we summarize the functions of extracellular ATP and adenosine in the context of tumor immunity. Second, we present an overview of the immunosuppressive and pro-angiogenic effects of extracellular adenosine. Third, we present experimental evidence that extracellular ATP and adenosine receptors are expressed by tumor cells and enhance tumor growth. Finally, we discuss recent studies, including our own work, which suggest that therapeutic approaches that promote ATP-mediated activation of inflammasomes, or inhibit the accumulation of tumor-derived extracellular adenosine, may constitute effective new means to induce anticancer activity.

Regression of murine lung tumors by the let-7 microRNA.

Abstract: MicroRNAs (miRNAs) have recently emerged as an important new class of cellular regulators that control various cellular processes and are implicated in human diseases, including cancer. Here, we show that loss of let-7 function enhances lung tumor formation in vivo, strongly supporting the hypothesis that let-7 is a tumor suppressor. Moreover, we report that exogenous delivery of let-7 to established tumors in mouse models of non-small-cell lung cancer (NSCLC) significantly reduces the tumor burden. These results demonstrate the therapeutic potential of let-7 in NSCLC and point to miRNA replacement therapy as a promising approach in cancer treatment.

ZEB1 represses E-cadherin and induces an EMT by recruiting the SWI/SNF chromatin-remodeling protein BRG1.

Abstract: Loss of E-cadherin is a key initial step in the transdifferentiation of epithelial cells to a mesenchymal phenotype, which occurs when tumor epithelial cells invade into surrounding tissues. Expression of the nuclear factor ZEB1 induces an epithelial-to-mesenchymal transition and confers a metastatic phenotype on carcinomas by repressing the E-cadherin gene at the transcriptional level. In this study, we show that ZEB1 interacts with the SWI/SNF chromatin-remodeling protein BRG1 to regulate E-cadherin independently of CtBP, its traditional co-repressor. Blocking the interaction between ZEB1 and BRG1 induces expression of E-cadherin and downregulation of the mesenchymal marker vimentin. ZEB1 and BRG1 colocalize in E-cadherin-negative cells from cancer lines and in the stroma of normal colon. Colocalization of ZEB1 and BRG1 in epithelial cells is only found in those de-differentiated cells characterized by nuclear β-catenin staining at the invasive edge of the tumor. Our results identify ZEB1/BRG1 as a new transcriptional mechanism regulating E-cadherin expression and epithelial-to-mesenchymal transdifferentiation that may be involved during the initial stages of tumor invasion.

HMGB1 release and redox regulates autophagy and apoptosis in cancer cells.

Abstract: The functional relationship and cross-regulation between autophagy and apoptosis is complex. In this study we show that the high-mobility group box 1 protein (HMGB1) is a redox-sensitive regulator of the balance between autophagy and apoptosis. In cancer cells, anticancer agents enhanced autophagy and apoptosis, as well as HMGB1 release. HMGB1 release may be a prosurvival signal for residual cells after various cytotoxic cancer treatments. Diminished HMGB1 by short hairpin RNA transfection or inhibition of HMGB1 release by ethyl pyruvate or other small molecules led predominantly to apoptosis and decreased autophagy in stressed cancer cells. In this setting, reducible HMGB1 binds to the receptor for advanced glycation end products (RAGEs), but not to Toll-like receptor 4, induces Beclin1-dependent autophagy and promotes tumor resistance to alkylators (melphalan), tubulin disrupting agents (paclitaxel), DNA crosslinkers (ultraviolet light) and DNA intercalators (oxaliplatin or adriamycin). On the contrary, oxidized HMGB1 increases the cytotoxicity of these agents and induces apoptosis mediated by the caspase-9/-3 intrinsic pathway. HMGB1 release, as well as its redox state, thus links autophagy and apoptosis, representing a suitable target when coupled with conventional tumor treatments.

Multiple microRNAs modulate p21Cip1/Waf1 expression by directly targeting its 3′ untranslated region

Abstract: Cyclin-dependent kinase inhibitor 1A (CDKN1A), also known as p21Cip1/Waf1, is a master downstream effector of tumor suppressors. In this study, we experimentally demonstrate through a high-throughput luciferase reporter screen that p21Cip1/Waf1 can be directly targeted by nearly 28 microRNAs (miRNAs). The results were further confirmed by a series of mutational analyses and luciferase reporter assays. These 28 miRNAs can substantially inhibit p21Cip1/Waf1 expression, predominantly at translational level. Many of these miRNAs were upregulated in cancers and might serve as modulators of oncogenesis. Furthermore, 8 of these 28 p21-regulating miRNAs are located in the chromosome 19 miRNA cluster, the largest miRNA gene cluster in humans, and they can clearly promote cell proliferation and cell-cycle progression in choriocarcinoma cells. In conclusion, our screening strategy provides an alternative approach to uncovering miRNA modulators of an individual mRNA, and it has identified multiple miRNAs that can suppress p21Cip1/Waf1 expression by directly targeting its 3' untranslated region.

Integrative molecular profiling of triple negative breast cancers identifies amplicon drivers and potential therapeutic targets.

Abstract: Triple negative breast cancers (TNBCs) have a relatively poor prognosis and cannot be effectively treated with current targeted therapies. We searched for genes that have the potential to be therapeutic targets by identifying genes consistently overexpressed when amplified. Fifty-six TNBCs were subjected to high-resolution microarray-based comparative genomic hybridization (aCGH), of which 24 were subjected to genome-wide gene expression analysis. TNBCs were genetically heterogeneous; no individual focal amplification was present at high frequency, although 78.6% of TNBCs harboured at least one focal amplification. Integration of aCGH and expression data revealed 40 genes significantly overexpressed when amplified, including the known oncogenes and potential therapeutic targets, FGFR2 (10q26.3), BUB3 (10q26.3), RAB20 (13q34), PKN1 (19p13.12) and NOTCH3 (19p13.12). We identified two TNBC cell lines with FGFR2 amplification, which both had constitutive activation of FGFR2. Amplified cell lines were highly sensitive to FGFR inhibitor PD173074, and to RNAi silencing of FGFR2. Treatment with PD173074 induced apoptosis resulting partly from inhibition of PI3K-AKT signalling. Independent validation using publicly available aCGH data sets revealed FGFR2 gene was amplified in 4% (6/165) of TNBC, but not in other subtypes (0/214, P=0.0065). Our analysis demonstrates that TNBCs are heterogeneous tumours with amplifications of FGFR2 in a subgroup of tumours.

CD24 + cells from hierarchically organized ovarian cancer are enriched in cancer stem cells

Abstract: Cancer stem cells (CSCs) have been identified in solid tumors and cancer cell lines In this study, we isolated a series of cancer cell clones, which were heterogeneous in growth rate, cell cycle distribution and expression profile of genes and proteins, from ovarian tumor specimens of a patient and identified a sub-population enriched for ovarian CSCs defined by CD24 phenotype Experiments in vitro demonstrated CD24(+) sub-population possessed stem cell-like characteristics of remaining quiescence and more chemoresistant compared with CD24(-) fraction, as well as a specific capacity for self-renewal and differentiation In addition, injection of 5 x 10(3) CD24(+) cells was able to form tumor xenografts in nude mice, whereas equal number of CD24(-) cells remained nontumorigenic We also found that CD24(+) cells expressed higher mRNA levels of some 'stemness' genes, including Nestin, beta-catenin, Bmi-1, Oct4, Oct3/4, Notch1 and Notch4 which were involved in modulating many functions of stem cells, and lower E-cadherin mRNA level than CD24(-) cells Altogether, these observations suggest human ovarian tumor cells are organized as a hierarchy and CD24 demarcates an ovarian cancer-initiating cell population These findings will have important clinical applications for developing effective therapeutic strategies to treat ovarian cancer

Cross talk between apoptosis and autophagy by caspase-mediated cleavage of Beclin 1

Abstract: Beclin 1 has a key role in the initiation of autophagy, a process of self-cannibalism in which cytoplasmic constituents are sequestered and targeted for lysosomal degradation. In a recent issue of Cell Death & Disease, Wirawan et al. report the significant finding that caspases can cleave Beclin 1, thereby destroying its pro-autophagic activity. Moreover, the C-terminal fragment of Beclin 1 that results from this cleavage acquires a new function and can amplify mitochondrion-mediated apoptosis. Of note, the BH3 domain of Beclin 1 remains within the N-terminal fragment, which has no detectable pro-apoptotic activity. These findings provide important insights into the molecular cross talk between autophagy and apoptosis.

Hypoxia-regulated microRNA-210 modulates mitochondrial function and decreases ISCU and COX10 expression

Abstract: The mechanisms of compromised mitochondrial function under various pathological conditions, including hypoxia, remain largely unknown. Recent studies have shown that microRNA-210 (miR-210) is induced by hypoxia under the regulation of hypoxia-inducible factor-1α and has an important role in cell survival under hypoxic microenvironment. Hence, we hypothesized that miR-210 has a role in regulating mitochondrial metabolism and investigated miR-210 effects on mitochondrial function in cancer cell lines under normal and hypoxic conditions. Our results demonstrate that miR-210 decreases mitochondrial function and upregulates the glycolysis, thus make cancer cells more sensitive to glycolysis inhibitor. miR-210 can also activate the generation of reactive oxygen species (ROS). ISCU (iron-sulfur cluster scaffold homolog) and COX10 (cytochrome c oxidase assembly protein), two important factors of the mitochondria electron transport chain and the tricarboxylic acid cycle have been identified as potential targets of miR-210. The unique means by which miR-210 regulates mitochondrial function reveals an miRNA-mediated link between microenvironmental stress, oxidative phosphorylation, ROS and iron homeostasis.

Membrane-bound mucins: the mechanistic basis for alterations in the growth and survival of cancer cells.

Abstract: Mucins (MUC) are high molecular weight O-linked glycoproteins whose primary functions are to hydrate, protect, and lubricate the epithelial luminal surfaces of the ducts within the human body. The MUC family is comprised of large secreted gel forming and transmembrane (TM) mucins. MUC1, MUC4, and MUC16 are the well-characterized TM mucins and have been shown to be aberrantly overexpressed in various malignancies including cystic fibrosis, asthma, and cancer. Recent studies have uncovered the unique roles of these mucins in the pathogenesis of cancer. These mucins possess specific domains that can make complex associations with various signaling pathways, impacting cell survival through alterations of cell growth, proliferation, death, and autophagy. The cytoplasmic domain of MUC1 serves as a scaffold for interaction with various signaling proteins. On the other hand, MUC4 mediates its effect by stabilizing and enhancing the activity of growth factor receptor ErbB2. MUC16, previously known as CA125, is a well-known serum marker for the diagnosis of ovarian cancer and has a key role in stimulation and dissemination of ovarian cancer cells by interacting with mesothelin and galectin. Therefore, herein we discuss the function and divergent mechanisms of MUC1, MUC4, and MUC16 in carcinogenesis in the context of alteration in cell growth and survival.

TGFβ mediated upregulation of hepatic miR-181b promotes hepatocarcinogenesis by targeting TIMP3

Abstract: To identify microRNAs (miRNAs) that may have a causal role in hepatocarcinogenesis, we used an animal model in which C57BL/6 mice fed choline-deficient and amino acid defined (CDAA) diet develop preneoplastic lesions at 65 weeks and hepatocellular carcinomas after 84 weeks. miRNA expression profiling showed significant upregulation of miR-181b and miR-181d in the livers of mice as early as 32 weeks that persisted at preneoplastic stage. The expression of tissue inhibitor of metalloprotease 3 (TIMP3), a tumor suppressor and a validated miR-181 target, was markedly suppressed in the livers of mice fed CDAA diet. Upregulation of hepatic transforming growth factor (TGF)β and its downstream mediators Smad 2, 3 and 4 and increase in phospho-Smad2 in the liver nuclear extract correlated with elevated miR-181b/d in mice fed CDAA diet. The levels of the precursor and mature miR-181b were augmented on exposure of hepatic cells to TGFβ and were significantly reduced by small interference RNA-mediated depletion of Smad4, showing the involvement of TGFβ signaling pathway in miR-181b expression. Ectopic expression and depletion of miR-181b showed that miR-181b enhanced matrix metallopeptidases (MMP)2 and MMP9 activity and promoted growth, clonogenic survival, migration and invasion of hepatocellular carcinoma (HCC) cells that could be reversed by modulating TIMP3 level. Further, depletion of miR-181b inhibited tumor growth of HCC cells in nude mice. miR-181b also enhanced resistance of HCC cells to the anticancer drug doxorubicin. On the basis of these results, we conclude that upregulation of miR-181b at early stages of feeding CDAA diet promotes hepatocarcinogenesis.

Nutrient sensor O-GlcNAc transferase regulates breast cancer tumorigenesis through targeting of the oncogenic transcription factor FoxM1.

Abstract: Cancer cells upregulate glycolysis, increasing glucose uptake to meet energy needs. A small fraction of a cell's glucose enters the hexosamine biosynthetic pathway (HBP), which regulates levels of O-linked beta-N-acetylglucosamine (O-GlcNAc), a carbohydrate posttranslational modification of diverse nuclear and cytosolic proteins. We discovered that breast cancer cells upregulate the HBP, including increased O-GlcNAcation and elevated expression of O-GlcNAc transferase (OGT), which is the enzyme catalyzing the addition of O-GlcNAc to proteins. Reduction of O-GlcNAcation through RNA interference of OGT in breast cancer cells leads to inhibition of tumor growth both in vitro and in vivo and is associated with decreased cell-cycle progression and increased expression of the cell-cycle inhibitor p27(Kip1). Elevation of p27(Kip1) was associated with decreased expression and activity of the oncogenic transcription factor FoxM1, a known regulator of p27(Kip1) stability through transcriptional control of Skp2. Reducing O-GlcNAc levels in breast cancer cells decreased levels of FoxM1 protein and caused a decrease in multiple FoxM1-specific targets, including Skp2. Moreover, reducing O-GlcNAcation decreased cancer cell invasion and was associated with the downregulation of matrix metalloproteinase-2, a known FoxM1 target. Finally, pharmacological inhibition of OGT in breast cancer cells had similar anti-growth and anti-invasion effects. These findings identify O-GlcNAc as a novel mechanism through which alterations in glucose metabolism regulate cancer growth and invasion and suggest that OGT may represent novel therapeutic targets for breast cancer.

New insights into apoptosis signaling by Apo2L/TRAIL

Abstract: Apoptosis ligand 2 tumor necrosis factor (TNF)-related apoptosis-inducing ligand (Apo2L/TRAIL) belongs to a small subset of proapoptotic protein ligands in the TNF superfamily. This subset, which also includes Fas ligand and TNF-α, can activate the extrinsic apoptotic cell death pathway on binding to cognate death receptors at the cell surface. Over the past 10 years, Apo2L/TRAIL has emerged as a promising candidate for cancer therapy, on the basis of its unique ability to trigger apoptosis in various types of cancer cells without significant toxicity toward normal cells. Herein, we review key advances in understanding the molecular events that control apoptosis signaling by Apo2L/TRAIL, which may aid in the development of cancer therapies based on the extrinsic apoptotic pathway.

Regulation of autophagy by ATF4 in response to severe hypoxia.

Abstract: Activating transcription factor 4 (ATF4) is a transcription factor induced under severe hypoxia and a component of the PERK pathway involved in the unfolded protein response (UPR), a process that protects cells from the negative consequences of endoplasmic reticulum (ER) stress. In this study, we have used small interfering RNA (siRNA) and microarray analysis to provide the first whole-genome analysis of genes regulated by ATF4 in cancer cells in response to severe and prolonged hypoxic stress. We show that ATF4 is required for ER stress and hypoxia-induced expansion of autophagy. MAP1LC3B (LC3B) is a key component of the autophagosomal membrane, and in this study we demonstrate that ATF4 facilitates autophagy through direct binding to a cyclic AMP response element binding site in the LC3B promoter, resulting in LC3B upregulation. Previously, we have shown that Bortezomib-induced ATF4 stabilization, which then upregulated LC3B expression and had a critical role in activating autophagy, protecting cells from Bortezomib-induced cell death. We also showed that severe hypoxia stabilizes ATF4. In this study, we demonstrate that severe hypoxia leads to ER stress and induces ATF4-dependent autophagy through LC3 as a survival mechanism. In summary, we show that ATF4 has a key role in the regulation of autophagy in response to ER stress and provide a direct mechanistic link between the UPR and the autophagic machinery.

Therapeutic CDK4/6 inhibition in breast cancer: Key mechanisms of response and failure

Abstract: A hallmark of cancer is the deregulation of cell-cycle machinery, ultimately facilitating aberrant proliferation that fuels tumorigenesis and disease progression. Particularly, in breast cancers, cyclin D1 has a crucial role in the development of disease. Recently, a highly specific inhibitor of CDK4/6 activity (PD-0332991) has been developed that may have efficacy in the treatment of breast cancer. To interrogate the utility of PD-0332991 in treating breast cancers, therapeutic response was evaluated on a panel of breast cancer cell lines. These analyses showed that the chronic loss of Rb is specifically associated with evolution to a CDK4/6-independent state and, ultimately, resistance to PD-0332991. However, to interrogate the functional consequence of Rb directly, knockdown experiments were performed in models that represent immortalized mammary epithelia and multiple subtypes of breast cancer. These studies showed a highly specific role for Rb in mediating the response to CDK4/6 inhibition that was dependent on transcriptional repression manifest through E2F, and the ability to attenuate CDK2 activity. Acquired resistance to PD-03322991 was specifically associated with attenuation of CDK2 inhibitors, indicating that redundancy in CDK functions represents a determinant of therapeutic failure. Despite these caveats, in specific models, PD-0332991 was a particularly effective therapy, which induced Rb-dependent cytostasis. Combined, these findings indicate the critical importance of fully understanding cell-cycle regulatory pathways in directing the utilization of CDK inhibitors in the clinic.

Activation of the hedgehog-signaling pathway in human cancer and the clinical implications

Abstract: The hedgehog pathway, initially discovered by two Nobel laureates Drs E Wieschaus and C Nusslein-Volhard in Drosophila, is a major regulator for cell differentiation, tissue polarity and cell proliferation. Studies from many laboratories reveal activation of this pathway in a variety of human cancer, including basal cell carcinomas (BCCs), medulloblastomas, leukemia, gastrointestinal, lung, ovarian, breast and prostate cancers. It is thus believed that targeted inhibition of hedgehog signaling may be effective in treatment and prevention of human cancer. Even more exciting is the discovery and synthesis of specific signaling antagonists for the hedgehog pathway, which have significant clinical implications in novel cancer therapeutics. In this review, we will summarize major advances in the last 2 years in our understanding of hedgehog signaling activation in human cancer, interactions between hedgehog signaling and other pathways in carcinogenesis, potential antagonists for hedgehog signaling inhibition and their clinical implications for human cancer treatment.

Proteasome inhibitors activate autophagy as a cytoprotective response in human prostate cancer cells.

Abstract: The ubiquitin-proteasome and lysosome-autophagy pathways are the two major intracellular protein degradation systems that work cooperatively to maintain homeostasis. Proteasome inhibitors (PIs) have clinical activity in hematological tumors, and inhibitors of autophagy are also being evaluated as potential antitumor therapies. In the current study we found that chemical proteasome inhibitors and siRNA-mediated knockdown of the proteasome's enzymatic subunits promoted autophagosome formation, stimulated autophagic flux, and upregulated expression of the autophagy specific genes (ATGs) (ATG5 and ATG7) in some human prostate cancer cells and immortalized mouse embryonic fibroblasts (MEFs). Upregulation of ATG5 and ATG7 only occurred in cells displaying PIs-induced phosphorylation of the eukaryotic translation initiation factor 2alpha (eIF2α), an important component of the unfolded protein responses. Furthermore, PIs did not induce autophagy or upregulate ATG5 in MEFs expressing a phosphorylation-deficient mutant form of eIF2α. Combined inhibition of autophagy and the proteasome induced an accumulation of intracellular protein aggregates reminiscent of neuronal inclusion bodies and caused more cancer cell death than blocking either degradation pathway alone. Overall, our data demonstrate that proteasome inhibition activates autophagy via a phospho-eIF2α-dependent mechanism to eliminate protein aggregates and alleviate proteotoxic stress.

Mir-30 reduction maintains self-renewal and inhibits apoptosis in breast tumor-initiating cells

Abstract: Accumulating evidence indicates that a sub-population of cancer cells with stem-like properties, termed tumor-initiating cells (T-ICs), exist in many different kinds of malignancies, which have a pivotal role in tumorigenesis, tumor progression, metastasis and post-treatment relapse. However, how the stem-like properties of T-ICs are regulated remains obscure. Our previous study showed that reduction of let-7 microRNA (miRNA) in breast tumor-initiating cells (BT-ICs) contributes to the maintenance of their self-renewal capacity and undifferentiated status. In this study we show the effect of mir-30 reduction on the stem-like features of BT-ICs. Similar to let-7, mir-30 is reduced in BT-ICs, and the protein level of Ubc9 (ubiquitin-conjugating enzyme 9) and ITGB3 (integrin beta3), the target genes of mir-30, is markedly upregulated. Enforced constitutive expression of mir-30 in BT-ICs inhibits their self-renewal capacity by reducing Ubc9, and induces apoptosis through silencing ITGB3. On the contrary, blocking the miRNA with a specific antisense oligonucleotide (ASO) in differentiated breast cancer cells revived their self-renewal capacity. Furthermore, ectopic expression of mir-30 in BT-IC xenografts reduces tumorigenesis and lung metastasis in nonobese diabetic/severe combined immunodeficient mice, whereas blocking mir-30 expression enhances tumorigenesis and metastasis. Together, our data suggest mir-30 as one of the important miRNAs in regulating the stem-like features of T-ICs.

Targeting mTOR: prospects for mTOR complex 2 inhibitors in cancer therapy.

Abstract: Small molecule inhibitors that selectively target cancer cells and not normal cells would be valuable anti-cancer therapeutics. The mammalian target of rapamycin complex 2 (mTORC2) is emerging as a promising candidate target for such an inhibitor. Recent studies in cancer biology indicate that mTORC2 activity is essential for the transformation and vitality of a number of cancer cell types, but in many normal cells, mTORC2 activity is less essential. These studies are intensifying interest in developing inhibitors that specifically target mTORC2. However, there are many open questions regarding the function and regulation of mTORC2 and its function in both normal and cancer cells. Here, we summarize exciting new research into the biology of mTORC2 signaling and highlight the current state and future prospects for mTOR-targeted therapy.

Ribosome biogenesis surveillance: probing the ribosomal protein-Mdm2-p53 pathway.

Abstract: The dynamic processes of cell growth and cell division remain under constant surveillance. As one of the primary 'gatekeepers' of the cell, p53 has a major role in sensing a variety of stressors to maintain cellular homeostasis. Growth is driven by new protein synthesis, a process that requires robust manufacture of ribosomes in the nucleolus. Ribosome biogenesis is a complex process comprising transcription, modification, and processing of ribosomal RNA, production of ribosomal proteins (RPs) and auxiliary factors, and coordinated assembly of ribonucleoprotein particles to produce mature ribosomes. As the major function of the nucleolus, ribosome biogenesis demands a considerable amount of resources and must be maintained in a coordinated manner to ensure fidelity of the process. Perturbations to many aspects of ribosome biogenesis are thought to contribute to 'nucleolar stress' and trigger a RP-Mdm2-p53 stress response pathway. In this review, we will clarify how disruption to three major components of ribosome biogenesis can trigger nucleolar stress and activate p53, thereby lending support to a RP-Mdm2-p53 ribosome biogenesis surveillance pathway.

Downregulation of microRNAs directs the EMT and invasive potential of anaplastic thyroid carcinomas.

Abstract: Anaplastic thyroid carcinomas (ATCs) arise from epithelial thyroid cells by mesenchymal de-/transdifferentiation and rapidly invade the adjacent tissue. Specific microRNA signatures were suggested to distinguish ATCs from normal thyroid tissue and other thyroid carcinomas of follicular origin. Whether distinct microRNA patterns correlate with de-/transdifferentiation and invasion of ATCs remained elusive. We identified two significantly decreased microRNA families that unambiguously distinguish ATCs from papillary and follicular thyroid carcinomas: miR-200 and miR-30. Expression of these microRNAs in mesenchymal ATC-derived cells reduced their invasive potential and induced mesenchymal-epithelial transition (MET) by regulating the expression of MET marker proteins. Supporting the role of transforming growth factor (TGF)beta signaling in modulating MET/epithelial-mesenchymal transition (EMT), expression of SMAD2 and TGFBR1, upregulated in most primary ATCs, was controlled by members of the miR-30 and/or miR-200 families in ATC-derived cells. Inhibition of TGFbeta receptor 1 (TGFBR1) in these cells induced MET and reduction of prometastatic miR-21, but caused an increase of the miR-200 family. These findings identify altered microRNA signatures as potent markers for ATCs that promote de-/transdifferentiation (EMT) and invasion of these neoplasias. Hence, TGFBR1 inhibition could have a significant potential for the treatment of ATCs and possibly other invasive tumors.

Cancer-associated IDH mutations: biomarker and therapeutic opportunities.

Abstract: The discovery of somatic mutations in the isocitrate dehydrogenase (IDH) enzymes through a genome-wide mutational analysis in glioblastoma represents a milestone event in cancer biology. The nature of the heterozygous, point mutations mapping to arginine residues involved in the substrate binding inspired several research teams to investigate their impact on the biochemical activity of these enzymes. Soon, it became clear that the mutations identified impaired the ability of IDH1 and IDH2 to catalyze the conversion of isocitrate to α-ketoglutarate (αKG), whereas conferring a gain of a novel enzymatic activity leading to the reduction of αKG to the metabolite D2-hydroxyglutarate (D-2HG). Across glioma as well as several hematologic malignancies, mutations in IDH1 and IDH2 have shown prognostic value. Several hypotheses implicating the elevated levels of D-2HG and tumorigenesis, and the therapeutic potential of targeting mutant IDH enzymes will be discussed.

Targeting of mRNAs by multiple miRNAs: the next step.

Abstract: Micro(mi)RNAs are small noncoding RNAs that regulate expression of the majority of the genes in the genome at either the messenger RNA (mRNA) level (by degrading mRNA) or the protein level (by blocking translation). miRNAs are thought to be components of vast regulatory networks. Currently, the field is focused primarily on identifying novel targets of individual miRNAs. This focus is about to undergo a dramatic change. In a new paper by Wu et al. (2010) it is experimentally confirmed that multiple miRNAs target the same gene, suggesting that it is the combination of all these activities that determines the expression of miRNA target genes. This study ushers in a new era of miRNA research that focuses on networks more than on individual connections between miRNA and strongly predicted targets.