bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1943: miR-148a overexpression reduces TRAIL resistance and tumorigenesis in NSCLC.
Authors: Joshi Pooja , Garofalo Michela , Croce Carlo M.
Published: August 2014

Non-small cell lung cancer (NSCLC) is the leading cause of death from cancer throughout the world. Although some advances have been made in lung cancer therapy, patient survival is still poor. Tumor-necrosis factor related apoptosis-inducing ligand (TRAIL) has been shown to induce apoptosis in malignant cells without inducing significant toxicity in normal cells. However, several carcinomas, including lung cancer, remain resistant to TRAIL. MicroRNAs (miRNAs) are small non-coding RNAs of ∼24 nt that can block mRNA translation and/or negatively regulate its stability. miRNAs are often aberrantly expressed in cancer and their function is linked to the regulation of oncogenes and/or tumor suppressor genes involved in cell signaling pathways. Several miRNAs have been implicated in increasing susceptibility or resistance to TRAIL-induced apoptosis by inhibiting key functional proteins.

In order to identify the miRNAs involved in TRAIL resistance in NSCLC, we evaluated the differential expression of miRNAs in H460 sensitive cells compared with H460R cells that have acquired TRAIL-resistance. MiR-148a was the most downregulated miR in the acquired resistant cells compared to the sensitive cells. Treatment of NSCLC cells with TRAIL after miR-148a overexpression decreased cell viability and increased apoptosis. Furthermore, miR-148a overexpression induced downregulation of matrix metalloproteinase 15 (MMP15) and Rho-associated kinase 1 (ROCK1) by targeting their 3’ untranslated regions as analysed by Western blot and luciferase reporter assay. Previous studies have shown that MMP15 acts as an anti-apoptotic factor involved in TRAIL resistance in cancer cells. Both, MMP15 and ROCK1 have been reported to be activators of tumor invasion and migration. Our preliminary functional studies using small interference RNA (siRNA) knockdowns of these genes in NSCLC cells support these data. A better understanding of the tumor-supressive role of miR-148a could be the key to developing an effective prognostic and therapeutic tool for treating NSCLC.

Citation Format: Pooja Joshi, Michela Garofalo, Carlo M. Croce. miR-148a overexpression reduces TRAIL resistance and tumorigenesis in NSCLC. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1943. doi:10.1158/1538-7445.AM2013-1943
Abstract 1952: Deregulated microRNA expression in the progression of salivary gland cancer.
Authors: Zheng Hao , Kiang Alan , Kuo Selena Z. , Rahimy Elham , Wang-Rodriguez Jessica , Ongkeko Weg M.
Published: August 2014

Salivary gland cancer constitutes one of the five main subsets of cancer in the head and neck region. Nonresectable malignant tumors demonstrate a low 20-year survival rate due to radioresistance and a high propensity for metastasis. The molecular mechanisms underlying pathogenesis of salivary cancer remains poorly elucidated, and thus necessitates further investigation. MicroRNAs (miRNA) are small noncoding RNAs that are predicted to regulate up to 30% of protein-encoding genes. Signature miRNA expression profiles have been identified in various malignancies, implicating miRNAs in the progression of human cancer. In order to better understand the role of miRNAs in the progression of salivary cancer, we profiled the miRNA expression levels of normal salivary tissue, benign salivary tumor, and salivary cancer. A total of 17 formalin-fixed paraffin-embedded salivary tissues, including 4 normal, 4 pleomorphic adenoma (PA), 3 squamous cell carcinoma (SCC), 3 mucoepidermoid carcinoma (ME) and 3 adenocarcinoma (AC), were collected and the endogenous expression of 95 miRNAs was analyzed by microarray. A large number of miRNAs were observed to be aberrantly expressed and generally down-regulated in salivary cancer and salivary tumor tissues with respect to normal salivary gland tissues. An unsupervised clustering and a student t-test were performed with a threshold p-value less than 0.05, resulting in the identification of 22 miRNAs differentially expressed at a statistically significant level. Eight candidate miRNAs were selected and further validated by RT-qPCR. The results suggest three miRNAs in SCC (miR-200a, let-7-family, and miR-192) and three miRNAs in AC (miR-107, miR-15b and miR-200a) to be highly involved in the carcinogenetic process of salivary cancers. Furthermore, the up-regulated expression pattern of miR-203 is indicative of a tumorigenetic function in salivary tumors. Interestingly, miR-200b was found to be consistently and significantly down regulated (p=0.01, 19.03 fold decrease) in the three types of salivary cancer under investigation. Recent studies have implicated miR-200b in EMT and invasion in cancer cells by directly targeting E-cadherin transcriptional repressors. The miRNAs identified in this study may serve as potential biomarkers and targets for future miRNA based therapy.

Citation Format: Hao Zheng, Alan Kiang, Selena Z. Kuo, Elham Rahimy, Jessica Wang-Rodriguez, Weg M. Ongkeko. Deregulated microRNA expression in the progression of salivary gland cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1952. doi:10.1158/1538-7445.AM2013-1952
Abstract 1953: Rapid quantitation of circulating microRNAs using a novel detection technology.
Authors: Duelli Dominik , Palma Jaime , Weldon Don , Doong Justin , Johnston Grace , Stacey Fellars , Hastings Michelle
Published: August 2014

The rapid implementation of quantifying microRNAs (miRNAs) in body fluids for diagnosis and prognosis could be improved by designing more rapid approaches. The current mainstream approach is quantitative reverse transcriptase PCR (qRT-PCR). Barriers for the applicability are lengthy time requirement for isolation and amplification of miRNAs, and polymerase inhibitors that copurify with miRNA, and compromise the accuracy of the approach. Here we provide an approach using a nano-particle technology that overcomes these limitations. This technology is based on the principle that a fluorophore-containing oligonucletide sequence is fluorescently silent unless displaced from a quencher by a specific substrate RNA. Once displacement occurs, the resulting fluorescence provides a measure of RNA concentration in the reaction. Our approach allows for the direct and rapid quantification of circulating miRNAs with minimal processing of blood plasma/serum and is done without the use of enzymes. Using circulating miRNAs with established roles in cancer and quality control, miR-16, miR-21 and miR-22 and miR-451, we identified optimal temperature, detergent and solvent-extraction parameters, to quantify accurately these miRNAs in plasma using a microplate fluorometer within an hour after plasma preparation. The stringency of our conditions can discriminate wild type from a single-basepair mismatch RNA, and is specific for mature miRNAs. Furthermore, this approach facilitates measurement of samples collected in heparin, an anticoagulant that severely inhibits polymerases used for qRT-PCR. This approach creates the possibility for rapid blood tests to quantify miRNA-associated events in real-time, and provides greater public access to miRNA studies by reducing the cost and time requirement of current accurate approaches of miRNA quantitation.

Citation Format: Dominik Duelli, Jaime Palma, Don Weldon, Justin Doong, Grace Johnston, Fellars Stacey, Michelle Hastings. Rapid quantitation of circulating microRNAs using a novel detection technology. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1953. doi:10.1158/1538-7445.AM2013-1953
Abstract 1957: MiR-200c can mediate either poor or good outcome in ovarian cancer depending on HuR localization in the cell.
Authors: Prislei Silvia , Martinelli Enrica , Mariani Marisa , Raspaglio Giuseppina , Sieber Steven , Ferrandina Gabriella , Shahabi Shohreh , Scambia Giovanni , Ferlini Cristiano
Published: August 2014

MicroRNAs (miRNAs) in solid malignancies can behave as predictors of either good or poor outcome. This is the case with members of the miR-200 family, which are the primary regulators of the epithelial to mesenchymal transition and have been reported to act as both oncogenes and tumor suppressors in independent studies. Expression of miR-200c was assessed in a panel of ovarian cancer cell lines with inherent or acquired drug-resistance. Luciferase reporter assay and ribonucleic-immuno-precipitation assay was used to in vitro characterise the complexes between miR-200c, HuR and class III β-tubulin (TUBB3). Nanofluidic technology and immunohistochemistry were used to analyze the expression of HuR, TUBB3 and miR-200c in 220 ovarian cancer patients. Using such approach, we demonstrated that overexpression of miR-200c in ovarian cancer resulted in poor or good outcome depending on interaction with the RNA binding protein HuR. When the localization of HuR was confined in the nucleus, high expression of miR-200c suppressed class III β-tubulin (TUBB3) expression and resulted in a good prognosis, whereas when HuR occurred in cytoplasm, the same miRNA enhanced TUBB3 expression and produced a poor outcome. Indeed, in an ovarian cancer cell line model, the miR-200c increased HuR binding on TUBB3 mRNA in stressing microenvironmental conditions. This study demonstrated how the prognostic power of a single factor can depend on interactions with additional factors. Through a multidimensional analysis of all components, the actual mechanism(s) underlying the aggressive biological behavior of ovarian cancer can be unraveled, and the prognosis of ovarian cancer patients can be rendered based on rational explanations.

Citation Format: Silvia Prislei, Enrica Martinelli, Marisa Mariani, Giuseppina Raspaglio, Steven Sieber, Gabriella Ferrandina, Shohreh Shahabi, Giovanni Scambia, Cristiano Ferlini. MiR-200c can mediate either poor or good outcome in ovarian cancer depending on HuR localization in the cell. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1957. doi:10.1158/1538-7445.AM2013-1957
Abstract 1958: Disregulation of TARBP-2 dependent miRNA maturation : A potential new therapeutic target in Ewing sarcoma..
Authors: De Vito Claudio , Riggi Nicolo , Cornaz Sandrine , Stamenkovic Ivan
Published: August 2014

We have recently shown that microRNAs (miRNAs) play essential roles is the generation of the cancer stem cell (CSC) phenotype in Ewing sarcoma familly tumors (ESFT). Here we compared the miRNA expression profile of ESFT CSCs to that of embryonic stem cells and CSCs from different tumor, which seem to be shared by these different cell types. Our data suggests that the mechanisms implicated in the miRNAs CSC signature rely on a reversible TARBP-2 dependent miRNA maturation defect. Restoration of TARBP2 activity either through chemical compounds or systemic delivery of synthetic forms of two of its targets, miRNA-143 or miRNA-145, inhibited ESFT CSC growth in vitro and in vivo. Taken together our observations provide new therapeutic strategies through restoration of normal miRNA expression profile, inhibiting CSC self-renewal and tumor maintenance.

Citation Format: Claudio De Vito, Nicolo Riggi, Sandrine Cornaz, Ivan Stamenkovic. Disregulation of TARBP-2 dependent miRNA maturation : A potential new therapeutic target in Ewing sarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1958. doi:10.1158/1538-7445.AM2013-1958
Abstract 1963: Prognosis-related miRNA expression profiles of neuroblastoma.
Authors: Ohira Miki , Isogai Eriko , Nakamura Yohko , Kamijo Takehiko , Nakagawara Akira
Published: August 2014

PURPOSE: Neuroblastoma (NB) is known to exhibit clinical heterogeneity and its accurate risk classification is one of the major challenges to improve the personalized therapy. We have been constructing the genomic/genetic profile-based risk classification system for NB. Recent studies indicated that miRNA expression levels are strongly correlated with patient prognosis of NB. To further improve our tumor stratification system by adding the miRNA classifier, we conducted miRNA expression profiling of favorable (F) and unfavorable (UF) NBs and searched for the prognosis-related miRNAs in our sample set.

METHODS: 48 primary samples (19 stage 1 or 2; 16 stage 3 or 4 without MYCN amplification; 13 stage 3 or 4 with MYCN amplification) were analyzed by miRNA microarray (Agilent G4470A). Statistical analysis was conducted by using the information of patient outcome at 3 years after diagnosis. Quantitative RT-PCR (qRT-PCR) was also conducted to validate the results in 101 primary tumors by using Taqman microRNA assays.

RESULTS: Among the 470 miRNAs analyzed, 79 miRNAs showed strong correlation to the prognosis (p<0.05): 17 and 62 miRNAs showed higher expressions in UF and F phenotypes, respectively. In the former miRNA group, 5 were located in 1q whose genomic copy number is frequently increased in UF type tumors. On the other hand, the latter includes 5, 6 and 9 miRNAs in 1p, 19q13 and 14q32, respectively, which are frequently decreased in copy numbers in UF type. In addition, miR-542-5p, whose expression has been reported as a prognostic marker by other groups, was also in the latter group in our dataset (p<0.001, 3.3-fold lower in UF). We further conducted qRT-PCR of these picked-up miRNAs with 101 independent NB samples for validation and found that higher expression of miR-181b and miR-149 showed significant correlation with poor and good prognosis of patients (both logrank p<0.001), respectively. In addition, intermediate-risk type of tumors (stage 3 or 4, no MYCN amplification, n=40) were also significantly classified into favorable or unfavorable by these two miRNA classifiers (p=0.021 and p=0.005, respectively). Interestingly, miR-149 has been reported that it has pro-apoptotic function by repressing Akt1 and E2F1 expressions in a neuroblastoma cell line.

CONCLUSION: The result suggested that miRNA expression profile exhibited strong correlation with NB prognosis, and integration of miRNA classifier with other markers could improve the prediction system for NB prognosis.

Citation Format: Miki Ohira, Eriko Isogai, Yohko Nakamura, Takehiko Kamijo, Akira Nakagawara. Prognosis-related miRNA expression profiles of neuroblastoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1963. doi:10.1158/1538-7445.AM2013-1963
Abstract 1969: The potential tumor-suppressive role of Krüppel-like factor 9 in colorectal cancer.
Authors: Brown Adam R. , Van Trang T. , Simmen Rosalia C.M. , Simmen Frank A.
Published: August 2014

Colorectal cancer (CRC) remains the third leading cause of cancer-related deaths in the United States. Recent attention has focused on several members of the Krüppel-like family (KLF) of transcription factors in pathogenesis of CRC. Published studies in mouse models have demonstrated that Krüppel-like Factors (KLF) 4 and 6 are important suppressors, while KLF5 is a promoter of CRC development and growth. Our and other laboratories have implicated KLF9 as another KLF with putative tumor-suppressive roles in colon and rectum. However, a causal role for KLF9 as a tumor suppressor has not been evaluated in vivo. In the present study, we generated C57BL/6J mice with two (wild-type, WT) and one (heterozygous, HET) Klf9 alleles within the ApcMin/+ model of intestinal cancer. At 16 weeks of age, male (by 2.5-fold, P=.0015) and female (by 1.5 fold, P=.002) Klf9 HET mice exhibited more colon tumors than WT (n=13-20 mice per gender/genotype). Further, HET females, but not males, showed increased (P=.011) colon tumor incidence compared to WT. Moreover, male HET exhibited greater numbers (P=.03) of large (>3mm diameter) colon tumors while corresponding female HET only showed a trend (P=.099), compared to WT. No significant difference was observed in the number of small intestine tumors in either males or females. Mean colon tumor size between genotypes of either sex was also comparable. Mice of both genotypes did not differ in body weights within specific female and male groups. Excised colon tumors and matched mucosa from male mice were embedded and subjected to immunohistochemistry. No significant differences in immunostaining intensity for β-Catenin were found in tumors nor in matched mucosa between the genotypes. A similar lack of differences in immunostaining for Lgr5, a surface marker for intestinal stem cells was noted in tumors or mucosa between the two genotypes. Since the ApcMin/+ model of intestinal cancer normally displays greater numbers of tumors in the small intestine than in the colon, results suggest a colon-specific contribution of KLF9 loss-of-expression to tumor development. Further, given that KLF9 is a unique thyroid hormone-inducible KLF family member, our results implicate the possible use of thyroid hormone or thyroid hormone receptor agonists in the prevention and/or treatment of CRC. Our collective findings provide early evidence for a tumor-suppressive role of KLF9 in the colon and rectum, the mechanism of which warrants further investigation. Supported by 1F31CA165665-01 and NIH RO1 CA136493.

Citation Format: Adam R. Brown, Trang T. Van, Rosalia C.M. Simmen, Frank A. Simmen. The potential tumor-suppressive role of Krüppel-like factor 9 in colorectal cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1969. doi:10.1158/1538-7445.AM2013-1969
Abstract 1971: Small C-terminal domain phosphatase 1 stabilizes PML to regulate the progression of renal clear cell carcinoma.
Authors: Lin Yu-Ching , Lu Li-Ting , Feng Xin-Hua , Chen Ruey-Hwa
Published: August 2014

The promyelocytic leukemia (PML) protein elicits pleiotropic tumor suppressive functions and is aberrantly degraded in multiple types of human cancers through poorly understood mechanisms. Previous study in our laboratory revealed that CDK1/2-mediated phosphorylation of PML Ser518 is required for targeting PML to Pin1 for prolylisomerization and subsequently to KLHL20-containing ubiquitin ligase complex for ubiquitination and proteasomal degradation. To identify the negative regulators of this PML degradation pathway, we employed a functional genomic approach to search for phosphatases that act on PML Ser518. This study identifies the small C-terminal domain phosphatase 1/2/3 (SCP1/2/3) as PML Ser518 phosphatases. Because these three phosphatases are of structural and functional similarities, we focus on the prototype of this family, SCP1. We show that SCP1 interacts with PML and reduces PML Ser518 phosphorylation levels both in vivo and in vitro. Through this dephosphorylation, SCP1 decreases Pin1-KLHL20 mediated-PML polyubiquitination and prolongs PML half life. Consequently, SCP1 increases the number of PML-nuclear bodies and elevates PML protein abundance in both normoxia and hypoxia conditions. At functional levels, SCP1 overexpression enhances multiple tumor suppressive effects, including promotion of apoptosis and inhibition of cell proliferation and migration. Importantly, all of these effects are reversed by PML knockdown. Clinically, SCP1 expression is downregulated in renal clear cell carcinoma (RCC) and this downregulation correlates with PML downregulation. Restoration of SCP1-mediated PML stabilization inhibits RCC proliferation, migration and transformation. Together, our study identifies SCP1 as a positive regulator of PML by dephosphorylation of PML Ser518 to antagonize the Pin1/KLHL20-dependent PML degradation pathway. Blockage of SCP1/PML axis is manifested in RCC and contributes to RCC progression.

Citation Format: Yu-Ching Lin, Li-Ting Lu, Xin-Hua Feng, Ruey-Hwa Chen. Small C-terminal domain phosphatase 1 stabilizes PML to regulate the progression of renal clear cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1971. doi:10.1158/1538-7445.AM2013-1971
Abstract 1973: Tumor suppressor p15Ink4b determines cell fate of hematopoietic progenitors: Implications for development of human blood disorders.
Authors: Humeniuk Rita , Rosu-Myles Michael , Wolff Linda
Published: August 2014

Red blood cells (RBCs) are a vital component of mammalian blood. Since they are short lived, they must be continuously replenished by erythropoiesis, a stepwise commitment of blood stem and progenitor cells to mature erythrocytes. The anemia due to the loss of RBC is a life threatening condition, often accompanying blood diseases such as leukemias and myelodysplastic syndromes (MDS). A striking 60-80% of these diseases have deleted or silenced expression of p15INK4B. An increased understanding of the factors that drive erythroid lineage commitment in progenitor cells is critical for developing new treatments for blood disorders. Previous examination of p15Ink4b knock-out mouse models, revealed skewing of hematopoietic progenitor differentiation towards myeloid lineage (granulocytes, macrophages). Here, we demonstrate a novel function for p15Ink4b in driving commitment to the erythroid lineage. Mice lacking p15Ink4b have lower numbers of primitive RBC progenitors and died shortly after induction of hemolytic anemia by phenylhydrazine injection. Expression of p15Ink4b in blood progenitors induced dynamic changes at the molecular level that rendered multi-lineage cells more permissive to erythroid commitment and less permissive to myeloid commitment. Noticeably, we found that p15Ink4b regulates a switch that controls the balance between myeloid and erythroid differentiation through activation of MEK/ERK signaling. In a time-coordinated manner expression of p15Ink4b induced rapid phosphorylation of MEK/ERK that led to rapid degradation of GATA-2 and activation of the GATA-1 transcription factors. Subsequently, the active GATA-1 executed lineage commitment through activation of the Erythropoietin receptor (EpoR), the “master regulator” of erythroid differentiation. The p15Ink4b mediated increase in GATA-1 expression, also resulted in decreased expression of the myeloid specific transcription factor PU.1, suppressing myeloid differentiation. In summary, we have defined a framework that determines how multipotent progenitors coordinate the balance between myeloid and erythroid differentiation. Central to this activity is p15Ink4b, which promotes erythroid fate while suppressing myeloid cell formation, a function that is particularly important in rapid RBC replenishment following stress. Our finding has implications not only for MDS and myeloid leukemia, where loss of tumor suppressor p15INK4B is a common event, but also for other forms of human refractory anemia.

Citation Format: Rita Humeniuk, Michael Rosu-Myles, Linda Wolff. Tumor suppressor p15Ink4b determines cell fate of hematopoietic progenitors: Implications for development of human blood disorders. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1973. doi:10.1158/1538-7445.AM2013-1973
Abstract 1977: The protein phosphatase 2 subunit PR48 is a novel melanoma tumor suppressor gene.
Authors: van Kempen Léon CL , Jarry Jonathan , Elchebly Mounib , Redpath Margaret , Edqvist Per-Henrík , Pontén Fredrik , Scolyer Richard , Schadendorf Dirk , Greenwood Celia M.T. , van den Oord Joost , Spatz Alan
Published: August 2014

Cutaneous melanoma cell proliferation is a strong prognostic factor for distant metastasis free survival (DMFS). In the current study we tested if aberrant expression of a regulator both DNA replication and cell cycle progression, i.e. the protein phosphatase 2A subunit PR48, could be a driver of melanoma proliferation and progression. Quantitative analysis of PR48 mRNA expression revealed a strong correlation between low levels of expression and poor DMFS in a multivariate analysis (n=49, p=0.0007). In line with this, no or low PR48 protein expression was associated with poor overall survival in three independent samples sets of melanoma (n=339, P<0.001), and reduced PR48 expression correlated with increased proliferation (P=0.0023). Functional analysis using over expression and knock down strategies in melanoma cell lines demonstrated that PR48 alters cdc6-cdt1 complex formation, regulates pRb phosphorylation status, and entry into the S-phase of the cycle progression. Ectopic PR48 expression decreased tumorigenicity of melanoma cells, and shRNA-mediated reduced PR48 expression increased tumor take and growth in nude mice. Our data demonstrate that PPP2R3B is a novel tumor suppressor gene in melanoma, whose loss of expression contributes increased proliferation and poor survival.

Citation Format: Léon CL van Kempen, Jonathan Jarry, Mounib Elchebly, Margaret Redpath, Per-Henrík Edqvist, Fredrik Pontén, Richard Scolyer, Dirk Schadendorf, Celia M.T. Greenwood, Joost van den Oord, Alan Spatz. The protein phosphatase 2 subunit PR48 is a novel melanoma tumor suppressor gene. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1977. doi:10.1158/1538-7445.AM2013-1977
Abstract 1978: Demonstrating a role for nuclear Adenomatous polyposis coli in intestinal cell differentiation.
Authors: Miller Matthew A. , Zeineldin Maged , Neufeld Kristi L.
Published: August 2014

Colon cancer is the second leading cause of cancer-related mortality in the United States, resulting in over 50,000 deaths annually. Approximately 80% of colon cancers begin with a mutation in the gene coding for the tumor suppressor protein Adenomatous Polyposis Coli, APC. APC protein can shuttle between the cytoplasm and the nucleus of cells, facilitated by two nuclear localization signals (NLS) and multiple nuclear export signals. To better understand functions of nuclear APC, our lab generated a “knock-in” mouse model with mutations introduced that inactivate both Apc NLS. These ApcmNLS/mNLS mice show a dramatic decrease in nuclear Apc. We previously showed increased proliferation and Wnt target gene expression in intestinal epithelial cells from ApcmNLS/mNLS mice, suggesting a role for nuclear APC in inhibition of proliferation and Wnt signaling. We also showed increased tumor number and size in ApcMin mice if they also harbor the ApcmNLS allele. Here we examine the role of nuclear Apc in intestinal epithelial differentiation and stem cell homeostasis. Paraffin-embedded tissue from the intestines of Apc+/+ and ApcmNLS/mNLS mice was sectioned and stained for markers of enterocytes (alkaline phosphatase) and goblet cells (alcian blue). Quantification of the positive cells indicated that ApcmNLS/mNLS mice had fewer differentiated enterocytes and goblet cells than their wild-type littermates. Intestines from ApcmNLS/mNLS mice were also stained for markers of quiescent and active stem cells, DCAMKL-1 and Lgr5 respectively. Quantification of the various stem cell populations will be described. Together, our data supports a role for nuclear Apc in promoting enterocyte and goblet cell differentiation, in suppression of tumors, and in inhibiting intestinal cell proliferation and Wnt signaling. Information gathered from analysis of ApcmNLS/mNLS mice will contribute to our understanding of the functions of nuclear APC in tumor suppression and ultimately the mechanism of intestinal tumorigenesis.

Citation Format: Matthew A. Miller, Maged Zeineldin, Kristi L. Neufeld. Demonstrating a role for nuclear Adenomatous polyposis coli in intestinal cell differentiation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1978. doi:10.1158/1538-7445.AM2013-1978
Abstract 1981: Ephrin B2 as a tumor suppressor in breast cancer.
Authors: Perez-Tenorio Gizeh , Stål Olle , Pasquale Elena B.
Published: August 2014

The EphB4 is a tyrosine kinase receptor with both tumor suppressor and promoting activities. The role of EphB4 in breast cancer is not fully elucidated. EphB-receptors transmit forward and reverse signals, which together are known as bidirectional signals. EphB4 reverse signaling, mediated by the ephrin-B2 ligand expressed in endothelial cells, causes some of the oncogenic properties of the receptor. In contrast, EphB4 forward signaling suppresses tumor growth. In breast cancers with high EphB4-receptor expression the forward signaling activity is low due to low expression of ephrin-B2. In fact, the EFNB2 gene, encoding ephrin-B2, is located on chromosome 13, which is lost in more than 30% of breast cancers. Our hypothesis was that ephrin-B2 re-expression in tumors could restore EphB4-forward signal inhibiting tumor growth. To examine the effects of EphB4-ephrin-B2 co-expression in vitro, we used lentiviral vectors to stably infect MCF7 cells with ephrin-B2. We found in vitro that ephrin-B2 expression caused cell detachment, inhibited cell proliferation; focus formation and cell motility, all characteristics of tumor suppression. Likewise, in patient materials we observed that high ephrin-B2 mRNA levels correlated with better distant recurrence-free survival. Interestingly, low ephrin-B2 expression identifies the group of worse prognosis among patients with high EphB4 and HER-2 expression in terms of distant recurrence and breast cancer survival. Taken together these results support our hypothesis and suggest that ephrin-B2 may be a tumor suppressor and a novel prognostic marker in breast cancer.

Citation Format: Gizeh Perez-Tenorio, Olle Stål, Elena B. Pasquale. Ephrin B2 as a tumor suppressor in breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1981. doi:10.1158/1538-7445.AM2013-1981
Abstract 1983: Functional analysis of cancer-associated mutations in human LATS1 and LATS2 tumor suppressors.
Authors: Yu Tian , Bachman John F. , Lai Zhi-Chun
Published: August 2014

Large Tumor Suppressor LATS1 and LATS2 (LATS1/2) are the key kinases of the hippo pathway, which regulates organ size and tumorigenesis in Drosophila and mammals in an evolutionary conserved pattern. Lats1 deficient mice develop soft tissue sarcomas, while disruption of Lats2 in mice and lats(wts) in Drosophila cause embryonic lethality. The Cancer Genome Project has identified 31 Lats1 and 21 Lats2 somatic mutations in the primary tumor tissues from a variety of human cancers, such as lung and ovarian cancer. However, the consequences of these mutations are still to be elucidated. In our study, evolutionary conservation analysis and structure remodeling predict 9 potentially damaging missense mutations, seven of which are located in the kinase domain. Immunoblotting demonstrates that one of the LATS2 mutants does not encode the protein product, whereas the other LATS1/2 mutants impair their phosphorylation, and attenuate their kinase activities on phosphorylating oncogene YAP. Furthermore, luciferase-based reporter assay shows that transcriptional coactivator activity of YAP is inhibited by wild type LATS1/2, but not by the mutants. In addition, in vivo transgenic expressions of LATS1/2 mutants result in less growth suppression in the Drosophila wing, when compared with the wild type. LATS1/2 mutants also fail to fully rescue the lethality causing by endogenous lats(wts) knockdown. Together, these results suggest that human LATS1 and LATS2 mutations deregulate oncogene YAP and highlight the crucial role of hippo pathway control in tumor suppression.

Citation Format: Tian Yu, John F. Bachman, Zhi-Chun Lai. Functional analysis of cancer-associated mutations in human LATS1 and LATS2 tumor suppressors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1983. doi:10.1158/1538-7445.AM2013-1983
Abstract 1985: ZNF783, a novel zinc finger protein has tumor suppressor function in prostate cancer.
Authors: Tbaishat Rana , Wang Songping , Kwabi-Addo Bernard
Published: August 2014

In the last decade hypermethylation of promoter regions has been accepted to be a prominent feature of human tumorigenesis by silencing tumor suppressor genes and other genes involved in cellular pathways. To study more in depth the involvement of aberrant methylation in prostate cancer tumorigenesis, we used a genome wide methylation array from

Illumina (Infunium) to identify novel hypermethylated genes that might contribute to prostate cancer progression. ZNF783 is one such candidate gene. ZNF783 is

Zinc-finger transcription factor that maps to human chromosome 7q33, a region implicated in prostate cancer tumorigenesis. We have observed significant hypermethylation of ZNF783 promoter CpG island in prostate cancer cell lines and prostate tumors when compared to match normal prostate tissue samples from radical prostatectomy patients. The treatment of prostate cancer cell lines with pharmacological demethylating agents restored ZNF783 gene expression indicating

a role for epigenetic DNA methylation in regulating ZNF783 expression in prostate cancer. The ZNF family of proteins has been identified to play essential roles in gene expression/regulation including inhibition of cell cycle proliferation, differentiation and apoptosis. The current study demonstrates a potential tumor suppressor role for ZNF783 in prostate cancer cells.

Citation Format: Rana Tbaishat, Songping Wang, Bernard Kwabi-Addo. ZNF783, a novel zinc finger protein has tumor suppressor function in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1985. doi:10.1158/1538-7445.AM2013-1985
Abstract 1986: Tumor suppressor PDCD4 inhibits NFκB dependent transcription in human glioblastoma cells by direct interaction with p65.
Authors: Hwang Soon-Kyung , Young Matthew R. , Colburn Nancy H.
Published: August 2014

PDCD4 was first described as a protein induced by apoptotic stimuli that acts as a tumor suppressor. PDCD4 plays an important role in regulating translation and transcription. Our previous research showed that overexpression of PDCD4 leads to decreased anchorage independent growth in glioblastoma-derived cell lines as well as decreased tumor growth in a glioblastoma xenograft model. Moreover, PDCD4 positively influences tumor necrosis factor-induced activation of the transcription factor NF-κB in inflammatory cells. However, the effect of PDCD4 on NF-κB transcriptional activity in cancer cells is still unknown. NF-κB acts as an oncogenic driver in many cancer sites. We studied the effect of PDCD4 on NF-κB-dependent transcriptional activity in malignant human glioblastoma cell lines by stably overexpressing PDCD4 in GBM U251 and LN229 cells. Stable Pdcd4 expression inhibits NF-κB transcriptional activation measured by a luciferase reporter in U251 and LN229 cells. The molecular mechanism by which PDCD4 inhibits NF-κB transcriptional activation does not involve inhibited expression of NF-κB p65 or p50 proteins. PDCD4 overexpression does not inhibit pathways upstream of NF-κB including the activation of IKKalpha and IKKbeta kinases. In addition, PDCD4 does not inhibit phosphorylation or degradation of IκBα, events needed for nuclear transport of p65 and p50. Overexpression of PDCD4 does, however, inhibit translocation of p65 but not p50 to the nucleus. Pdcd4 protein interacts directly with p65 protein as shown by co-immunoprecipitation and confocal imaging. These results suggest that PDCD4 can significantly inhibit NF-κB activity in glioblastoma cells by a mechanism that involves direct protein-protein interaction rather than the expected mRNA-selective translational inhibition. These findings may provide novel opportunities for NF-κB targeted interventions to prevent or treat cancer.

Citation Format: Soon-Kyung Hwang, Matthew R. Young, Nancy H. Colburn. Tumor suppressor PDCD4 inhibits NFκB dependent transcription in human glioblastoma cells by direct interaction with p65. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1986. doi:10.1158/1538-7445.AM2013-1986
Abstract 1988: Divergent control of Cav-1 expression in non-cancerous Li-Fraumeni Syndrome and human cancer cell lines.
Authors: Sherif Zaki A. , Sultan Ahmed S.
Published: August 2014

Li-Fraumeni Syndrome (LFS) is primarily characterized by development of tumors exhibiting germ-line mutations in the p53 gene. Cell lines developed from patients of a LFS family have decreased p53 activity as evidenced by the absence of apoptosis upon etoposide treatment. To test our hypothesis that changes in gene expression beyond p53 per se are contributing to the development of tumors, we compared gene expression in non-cancerous skin fibroblasts of LFS-affected (p53 heterozygous) versus non-affected (p53 wild-type homozygous) family members. Expression analysis showed that several genes were differentially regulated in the p53 homozygous and heterozygous cell lines. We were particularly intrigued by the decreased expression (∼88%) of a putative tumor suppressor protein, caveolin-1 (Cav-1), in the p53-mutant cells. Decreased expression of Cav-1 was also seen in both p53-knockout and p21-knockout HTC116 cells suggesting that p53 controls Cav-1 expression through p21 and leading to the speculation that p53, Cav-1 and p21may be part of a positive auto-regulatory feedback loop. The direct relationship between p53 and Cav-1 was also tested with HeLa cells (containing inactive p53), which expressed a significantly lower Cav-1 protein. A panel of nonfunctional and p53-deficient colon and epithelial breast cancer cell lines showed undetectable expression of Cav-1 supporting the role of p53 in the control of Cav-1. However, in two aggressively metastasizing breast cancer cell lines, Cav-1 was strongly expressed suggesting a possible role in tumor metastasis. Thus, there is a divergent control of Cav-1 expression as evidenced in non-cancerous Li-Fraumeni Syndrome and some aggressive human cancer cell lines.

Citation Format: Zaki A. Sherif, Ahmed S. Sultan. Divergent control of Cav-1 expression in non-cancerous Li-Fraumeni Syndrome and human cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1988. doi:10.1158/1538-7445.AM2013-1988
Abstract 1989: ASPP2 enhances DRAM induced autophagy to promote apoptosis in HepG2 cells.
Authors: Liu Kai , Wen Tao , Yin Ming Ji , Qiao Xin Lu , Shi Ying , Li Li , Liu Jie Dao , Zhang Chao , Li Ning , Lopez Charles , Chen Xi De
Published: August 2014

Background and Aims: ASPP2 (Apoptosis Stimulating Protein 2 of p53) binds to p53 and enhances p53 pro-apoptotic function. Autophagy may play a role in this process although the mechanisms remain to be clarified. Our aims were to explore the mechanisms by which the ASPP2-p53 pathway could promote apoptosis and autophagy in hepatocellular carcinoma cells in order to identify novel therapeutic targets in this disease.

Methods: HepG2 cells were transfected with GFP-LC3 plasmid and then damaged with methyl methanesulfonate (MMS). Apoptosis was detected by cleavage of CK18 and PARP by immunocytochemical staining and Western blot. Real-time PCR and immunoblot were used to analyze changes in DRAM and LC3I/II expression. DRAM siRNA was used to inhibit DRAM function.

Results: After MMS exposure for 24 hours, we identified more M30 positive cells and p85 formation in HepG2 cells compared to control cells. When HepG2 cells were transfected with a GFP-LC3 expression plasmid and then treated by MMS, we found that MMS treatment induced LC3 staining punctate structures, a marker for autophagy. Moreover, enforced expression of ASPP2 using adenovirus infection led to more autophagy and apoptosis in HepG2 after MMS treatment, under conditions where ASPP2 expressing adenovirus infection alone could not induce apoptosis and autophagy. Using real time PCR, we identified that MMS treatment could induce DRAM expression and that ASPP2 overexpression prior to MMS could further enhance DRAM expression compared to MMS alone. Immunoblot assay demonstrated that ASPP2 expression increased MMS-induced DRAM expression, LC3 II and p85 formation. Importantly, a DRAM specific siRNA blocked autophagy and apoptosis in HepG2 cells under these conditions when compared to a control siRNA.

Conclusions: Our results suggest that ASPP2 can enhance MMS induced apoptosis via p53-dependent DRAM-mediated autophagy. Since DRAM is an important component of p53-mediated autophagy, our results demonstrate a new role for ASPP2 in this pathway. These findings open new avenues for investigation into the mechanisms of autophagy-based therapy in hepatocellular cancer (Supported by National Natural Science Foundation of China (30870853,30770742 and 30910103915), Beijing Municipal Natural Science Foundation (7092045 and 7101005) and Basic-Clinical Collaborative Research Fund, CCMU (12JL-L05)).

Citation Format: Kai Liu, Tao Wen, Ming Ji Yin, Xin Lu Qiao, Ying Shi, Li Li, Jie Dao Liu, Chao Zhang, Ning Li, Charles Lopez, Xi De Chen. ASPP2 enhances DRAM induced autophagy to promote apoptosis in HepG2 cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1989. doi:10.1158/1538-7445.AM2013-1989
Abstract 1990: Loss of Olfactomedin 4 gene leads to neoplastic progression in the murine prostate.
Authors: Li Hongzhen , Liu Wenli , Zhu Jianqiong , Chen Weiping , Deng Chu-Xia , Rodgers Griffin P.
Published: August 2014

The murine olfactomedin 4 gene (Olfm4) encodes an olfactomedin-related glycoprotein whose physiological and pathological functions in the prostate are largely unknown. We have observed that homozygous Olfm4 mutant mice develop prostatic intra epithelial neoplisa (PIN), high grade PIN, adenocarcinoma and metastatic tumor in an age-dependent manner. We also found that increased proliferation of prostate epithelial cells in Olfm4-null mice. In this report, we studied the mechanisms underlying neoplastic progression of Olfm4-deficient mice prostate. We have performed microarray analysis to compare Olfm4 knockout and wild-type mice prostate at 3 and 15 months of ages. A total of 187 genes were found commonly changed in both 3 and 15 months, whereas 894 and 1090 genes were significantly changed in 3 and 15 months respectively. Enrichment Gene Go Diseases analysis showed prostatic neoplasms was at the top of 10 listed diseases. We found that 95 genes for neoplastic procession markers were significantly changed in the prostate of Olfm4-deficient mice. Among the genes, 39 genes were up-regulated and 37 genes were down regulated more than 1.5-fold. The qRT-PCR confirmed consistently higher expression of Ets1, Vav3, Shh, Sdc2, and Mmp19, and down-regulation of the genes Rnasel and Psca in Olfm4 knockout mice compared with Olfm4 wild-type mice. We also confirmed expression of Ets1 and Mmp19 by using western blot and immunohistochemistry analysis.

Interestingly, in the GEO prostate cancer database, which compares tumor to normal tissue and metastatic tumor to normal tissue, expression of OLFM4 and RNASEL were down-regulated, in contrast, expression of ETS1 and MMP19 were up-regulated. Therefore, the gene expression profiles in the GEO database confirmed and extended Ets1, Mmp19 and Rnasel in Olfm4 knockout mice. Our finding indicates that loss of OLFM4 leads to progression of prostate cancer.

Citation Format: Hongzhen Li, Wenli Liu, Jianqiong Zhu, Weiping Chen, Chu-Xia Deng, Griffin P. Rodgers. Loss of Olfactomedin 4 gene leads to neoplastic progression in the murine prostate. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1990. doi:10.1158/1538-7445.AM2013-1990
Abstract 1991: PDCD4 de-repression of Bcl-xl IRES-mediated translation leads to enhanced chemo-resistance of glioblastoma multiforme tumors.
Authors: Liwak Urszula , Jordan Lindsay E. , Roncaroli Federico , Holcik Martin
Published: August 2014

Glioblastoma multiforme (GBM) is the most prevalent form of tumor of the central nervous system with an average survival time of less than one year. Given the location and characteristics of GBM only limited treatment options are available, thus understanding the mechanisms of GBM formation or progression may lead to the development of novel treatment options. Strikingly, a common feature of GBM is the loss of expression of the tumor suppressor programmed cell death 4 (PDCD4) which correlates with adverse outcomes for patients in response to chemo- or radiation therapy. Through both transcriptional and translational regulation, PDCD4 has been shown to regulate many proteins within the cell, although the precise mechanism of target selection is not known. We have hypothesised that silencing of PDCD4 in glioblastomas leads to an altered expression of apoptosis-regulating proteins resulting in the suppression of apoptotic signals and enhanced chemo-resistance. Indeed, we show that low levels of PDCD4 correlate with an increase in the anti-apoptotic Bcl-xL protein. Importantly, Bcl-xL is regulated at the level of protein synthesis through the internal ribosome entry site (IRES) in its 5’ untranslated region (UTR). We further show that PDCD4 regulates the activity of the Bcl-xL IRES, and is therefore an IRES specific translational regulator. Importantly, reintroduction of PDCD4 into glioblastoma cells or directly inhibiting Bcl-xL with the chemical inhibitor ABT-737 sensitizes these cells to doxorubicin, thus enhancing cell death. In summary, our work provides a novel role for PDCD4 as a translational regulator of Bcl-xL as well as a potential therapeutic option for the treatment of GBM.

Citation Format: Urszula Liwak, Lindsay E. Jordan, Federico Roncaroli, Martin Holcik. PDCD4 de-repression of Bcl-xl IRES-mediated translation leads to enhanced chemo-resistance of glioblastoma multiforme tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1991. doi:10.1158/1538-7445.AM2013-1991