bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2033: Regulatory role of miR-203 in prostate cancer
Authors: Saini Sharanjot , Majid Shahana , Deng Guoren , Yamamura Soichiro , Ueno Koji , Chen Yi , Zaman Saif , Dahiya Rajvir
Published: February 2011

MicroRNAs (miRNAs) constitute an evolutionarily conserved class of small non-coding RNAs that primarily regulate gene expression at the post-transcriptional level via sequence-specific interaction with the 3’-untranslated regions (UTRs) of cognate mRNA targets. It is being increasingly recognized that miRNAs play roles in the establishment and progression of human malignancies. Several recent studies have shown that microRNAs are significantly altered in prostate cancer. Here we explored the role of miR-203 in prostate cancer progression and metastasis. RT-PCR analysis of prostate carcinoma cell lines showed that miR-203 expression is attenuated in prostate cancer. This attenuation was validated by in situ hybridization analysis of miR-203 in clinical specimens. To evaluate the functional significance of miR-203, miR-203 was reintroduced in prostate cancer cell lines followed by functional assays. Overexpression of miR-203 in prostate cancer cell line leads to decreases in cell growth, clonability, invasiveness and migratory properties. miR-203 reexpression in prostate carcinoma cell line induces cell cycle arrest at the G0-G1 phase of the cell cycle with a concomitant alteration of cell cycle regulators such as p21, p27, cyclin E and cdk2. Also, miR-203 transfection leads to a significant increase in apoptosis in prostate cancer cells pointing to the pleiotropic roles of this miR in prostate cancer. Further, analysis of putative miR-203 targets showed that miR-203 regulates the expression of survivin, a small inhibitor of apoptosis (IAP) protein that plays a key role in prostate cancer progression. In conclusion, our data suggests a crucial tumor suppressive role for miR-203 in prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2033.
Abstract 2034: Tumor suppressive activity of Smad3 by regulation of microRNAs in gastric cancer
Authors: Kim Jooyoung , Park Dong-Kyun , Ahn Sung-Min , Cha Jiyoung , Hong Suntaek
Published: February 2011

Loss of the tumor suppressive effect of transforming growth factor-β (TGF-β) has been commonly found at later stages in carcinogenic progression. It was recently found that Smad3 expression is lost in more than a third of cancers and the introduction of Smad3 into Smad3 null gastric cancer cell line restores TGF-β-mediated growth inhibitory and tumor suppressive activity. Interestingly, expression of Smad3 induced the expression of E-cadherin. Since there is accumulating evidence that microRNAs (miRNAs) regulate transcriptional repressors of E-cadherin such as ZEB1 and ZEB2, thus affecting the expression of E-cadherin, we aimed to compare miRNA profiles of Smad3-negative gastric cell line (SNU484-LPCX) and Smad3-overexpressed one (SNU484-Smad3). To investigate global changes of miRNA profiles associated with Smad3 restoration, we applied massively parallel sequencing (MPS) to miRNA profiling. Using this approach, we identified more than 150 known miRNAs, and at least 10 miRNAs including miR200 show statistical significance in expression between two cell lines. To further confirm these findings, we applied qRT-PCR analysis of selected miRNAs and mRNAs. Using Smad3 specific siRNAs, we also proved that Smad3 is involved in the E-cadherin expression through the regulation of specific miRNAs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2034.
Abstract 2036: MicroRNA-137 reverse the malignant phenotypes of colorectal cancer cells by targeting small GTPase Cdc42
Authors: BI FENG , Liu Ming , Lang Nan , Tang Qiuling
Published: February 2011

MicroRNAs have emerged as post-transcriptional regulators that are critically involved in the pathogenesis of a number of human cancers. Cdc42, one of the best characterized members of the Rho GTPase family, is found to be up-regulated in several types of human tumors, and has been implicated in cancer initiation and progression. In the present studies, we have identified miR-137 as a potential regulator of Cdc42 expression. A bioinformatics search revealed a target-site for miR-137 within the Cdc42 3’ UTR at nt 792-798, which is highly conserved across different species. Expression of miR-137 in colorectal cancer cell lines was found inversely correlated with Cdc42 expression. MiR-137 could significantly suppress Cdc42 3’ UTR luciferase-reporter activity, while this effect was not detectable when the putative target-site for miR-137 was mutated. Consistent with the results of reporter assay, ectopic expression of miR-137 reduced both mRNA and protein expression levels of Cdc42 and mimicked the effect of Cdc42 knockdown in inhibiting proliferation, inducing G1 cell cycle arrest, and blocking invasion of the colorectal cancer cells. Furthermore, expression of miR-137 suppressed the immediate downstream effector of Cdc42, PAK signaling in colorectal cancer cell lines. Our results suggest that miR-137 may have a tumor suppressor function by directly targeting the Rho GTPase Cdc42 to inhibit the proliferation and invasion activities of colorectal cancer cells. They raise an interesting possibility that Cdc42 activity and function can be controlled by microRNAs in addition to the classic regulators such as guanine nucleotide exchange factors and GTPase-activating proteins.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2036.
Abstract 2042: Aberrant expression of miR-21 triggers malignancy of human gliomas by disrupting negative feedback regulator, Spry2
Authors: Kim Youn-Jae , Kwak Hee-Jin , Park Jong Bae , Lee Seung-Hoon
Published: February 2011

The malignant progression of glioma is accompanied by diffuse infiltration. Recent evidences suggest the significant role of microRNAs in cancer cell migration and invasion. Therefore, we hypothesized that deregulation of microRNA may be important for the malignant progression of glioma. We found that aberrant expression of miR-21 is responsible for glioma invasion by disrupting the negative feedback circuit of Ras/MAPK signaling, which is mediated by Spry2. Upregulation of miR-21 was triggered by tumor microenvironmental factors such as hyaluronan and growth factors in glioma cells lacking functional PTEN, but not wild-type PTEN-harboring glioma cells. Consistently with these in vitro results, Spry2 protein levels were significantly decreased in 79.7% of invasive WHO grade II-IV human glioma tissues as well as cancer stem cell-derived tumor models, but not in non-invasive grade I and normal tissues. The Spry2 protein levels were not correlated with their mRNA levels, although they were inversely correlated with miR-21. Taken together, these results suggest that the post-transcriptional regulation of Spry2 by miR-21 plays an essential role on the malignant progression of human gliomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2042.
Abstract 2056: MicroRNA-126 inhibits SOX2 expression in gastric cancer cells
Authors: Otsubo Takeshi , Akiyama Yoshimitsu , Hashimoto Yutaka , Yuasa Yasuhito
Published: February 2011

A transcription factor SOX2 is essential for embryonic development and play critical roles in the maintenance of embryonic stem cell's pluripotency. We previously reported that SOX2 plays important roles in growth inhibition through cell cycle arrest and apoptosis, and SOX2 expression was frequently down-regulated in gastric cancers, some of which were due to aberrant DNA methylation. However, the mechanisms underlying loss of SOX2 expression and its target genes involved in gastric carcinogenesis remain largely unknown. In this study, we initially assessed whether microRNAs, small noncoding RNAs, control SOX2 expression in gastric cancers. In silico analysis revealed that microRNA-126 (miR-126) has two predictive target sites in the 3′-UTR of the SOX2 mRNA. Gain- and loss-of function experiments demonstrated that miR-126 inhibited SOX2 expression in gastric cancer cell lines. Luciferase reporter assays with a pGL4-SOX2 3′-UTR vector revealed that miR-126 directly inhibits SOX2 expression by targeting two binding sites in the 3′-UTR of the SOX2 mRNA independently. Some primary gastric cancer tissues with low expression of SOX2 exhibited markedly high levels of miR-126 expression compared with paired non-cancerous mucosae. To further understand the potential effects of miR-126-induced down-regulation of SOX2 on gastric carcinogenesis, we next attempted to identify the downstream target genes of SOX2 in gastric cancer cells. The microarray analysis after SOX2 over-expression in a gastric cancer cell line showed that 437 genes were up-regulated (>2-fold) and 447 genes were down-regulated (<0.5-fold), some of which were cancer-related genes. Taken together, we identified miR-126 as a novel microRNA which inhibits SOX2 expression in gastric cancer cells, and suggest that aberrant miR-126 expression and consequent SOX2 down-regulation may contribute to gastric carcinogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2056.
Abstract 2057: Cigarette smoke enhances pulmonary carcinogenesis via downregulation of has-miR-487b in respiratory epithelial cells
Authors: Xi Sichuan , Yang Maocheng , Tao Yong-Guang , Shan Jiqui , Xu Hong , Zhang Mary , Schrump David S.
Published: February 2011

Limited information is available regarding the mechanisms by which miRNA alternations contribute to initiation and progression of lung cancer. In order to examine this issue, array techniques were used to assess miRNA expression profiles in short term normal human bronchial epithelia (NHBE) and small airway epithelial cells (SAEC), as well as A549 and Calu-6 lung cancer cells cultured in normal media (NM) with or without cigarette smoke condensate (CSC). Under relevant exposure conditions, CSC consistently mediated a 3.4 − 5.5 fold decrease in has-miR-487b expression in cultured normal as well as lung cancer cells. Quantitative RT-PCR experiments demonstrated that expression levels of has-miR-487b were significantly downregulated (2.9 − 21 fold) in a panel of resected lung cancers relative to adjacent normal tissues. Software-guided analysis revealed numerous potential targets for has-miR-487b including H2AZ, encoding a histone variant modulating expression of developmentally regulated genes as well as SUZ12, which encodes a component of polycomb repressor complexes implicated in aberrant silencing of tumor suppressor genes such as Dickkopf-1 (Dkk-1) in lung cancer cells. Quantitative RT-PCR experiments revealed that constitutive over-expression of has-miR-487b mediated 3-35 fold reductions in H2AZ and SUZ12 expression in SAEC, A549, and Calu-6 cells. RNA cross-link immunoprecipitation (CLIP) experiments confirmed direct interference of has-miR-487b with H2AZ and SUZ12 transcripts. Overexpression of has-miR-487b enhanced sensitivity to cisplatin in lung cancer cells. Collectively, these data suggest that CSC-mediated downregulation of has-miR-487b promotes pulmonary carcinogenesis, in part, via activation of genes encoding proteins implicated in epigenetic regulation of stem cell gene expression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2057.
Abstract 2061: Vitamin D-regulates microRNAs in primary cultures of normal prostatic epithelial cells
Authors: Antonio Angeline , Vaishnav Avani , Nonn Larisa
Published: February 2011

Recent discoveries in cancer find that endogenous non-coding RNAs, microRNAs (miRNA), bind to the 3’ untranslated region of target mRNAs and function to translationally repress or decrease mRNA stability. Additionally, a number of studies show that aberrant miRNA expression is linked to various cancers, such as prostate cancer. Cancer-related changes in miRNA levels may result in either repression of tumor suppressor genes and/or activation of oncogenes. MiRNAs pose a potential role as targets in the treatment of cancers. In prostate cancer research, studies suggest a role of vitamin D as a chemopreventive agent. Vitamin D inhibits the growth and differentiation of malignant prostate cells and tumors. Studies show that a reduction in serum levels of vitamin D metabolites is linked to PC occurrence. Research from our laboratory shows that vitamin D treatment alters mRNA stability. This suggests a potential role for miRNAs in the vitamin D prevention of prostate cancer. We hypothesize that vitamin D's prevention of prostate cancer involves up-regulating tumor suppressive miRNAs and down-regulating oncogenic miRNAs. Using normal human prostatic epithelial cells as a model, we investigated the miRNA expression profile changes in cells that were treated with 1,25-dihydroxyvitamin D for time points ranging from 30 min to 24 hours. Our results demonstrated that approximately 50% of the miRNAs are expressed in the normal epithelial cells. A smaller percentage of miRNAs, approximately 10%, were changed by vitamin D treatment. We found that vitamin D down-regulated a small number of miRNAs (<0.5 fold) commonly up-regulated in cancer (miR-181d, miR-378) and up-regulated miRNAs that are down-regulated in cancer (miR-29a, miR-16). Of the miRNAs altered by vitamin D, we observed an interesting down-regulation of the let-7 family of tumor suppressor miRNAs. Six of the seven let-7 family members present on the array (let-7 a-g) were down-regulated across the time points with four of the let-7 miRNAs downregulated as early as 30 minutes. This is a somewhat discrepant finding since let-7 is known as a tumor suppressor miRNA that is decreased in many cancers and we know that vitamin D is a cancer preventive agent. Further studies will determine the role of vitamin D on let-7 in the prostatic epithelial cells and the implications in cancer. Ongoing research will also determine the mechanism of vitamin D regulation of miRNAs to help further understand the chemopreventive role of vitamin D in prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2061.
Abstract 2064: Down-regulation of miR-31 is associated with metastatic potential of human and canine osteosarcoma
Authors: Melkamu Tamene , Carlson Daniel F. , Farese James P. , Kassie Fekadu
Published: February 2011

Osteosarcoma is the most common primary bone tumor in humans and dogs. The primary cause for the poor survival of patients with osteosarcoma in both species is the emergence of metastases in organs distant from the primary tumor, especially in the lung. Therefore, identification of molecular determinants of osteosarcoma metastasis is highly warranted. In order to identify microRNAs (miRNAs) that regulate osteosarcoma metastasis, we examined, using qRT-PCR, levels of 10 miRNAs commonly associated with cancer metastasis (miR-10b, miR-146a, miR182, miR183, miR-206, miR-335, miR373, miR-520c-5p, miR-21 and miR-31) in total RNA samples prepared from canine parental osteosarcoma (POS) and highly metastatic POS (HMPOS) cells. Of the 10 miRNAs, only miR-31 showed differential expression between the two cell lines (reduced by 7.8-fold in HMPOS cells versus POS cells). Exposure of HMPOS cells to various chemopreventive agents (myo-inositol, indole-3-carbinol, diindolylmethane, deguelin, phenethyl isothiocyanate, cisplatin, and aza-2′-deoxycytidine) increased the level of miR-31 at least by two-fold, the most potent agents being deguelin and phenethyl isothiocyanate. Subsequently, we assessed the expression of miR-31 in additional canine osteosarcoma cell lines (three cell lines established from primary tumors and three cell lines developed from pulmonary metastases) and human osteosarcoma cell lines differing in metastatic potential (parental SaOS2, low- (LM2), and high- (LM7) metastatic cells). miR-31 levels decreased by 18-fold in canine cell lines derived from pulmonary metastatic tumors relative to that obtained from primary tumors and by 2-fold and 3-fold in LM2 and LM7 cells, respectively, compared to the expression in parental Saos-2 cells. Also, analysis of 3 primary human osteosarcoma tissues and 3 pulmonary metastatic tissues revealed a 13-fold reduction of miR-31 in metastatic tissues relative to primary tumor tissues. In preliminary experiments where attempts were made to identify potential targets of miR-31, exogenous over-expression of a synthetic miR-31 cassette in HMPOS cells negatively regulated a tight junction protein claudin-2, which is incriminated in cancer metastasis. In summary, our studies indicate that miR-31 is a promising marker for the metastatic potential of osteosarcoma and for disease prognosis. Moreover, reactivation of silenced miR-31 by chemopreventive/chemotherapeutic agents could reverse osteosarcoma metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2064.
Abstract 2065: DDX20 deficiency enhances NF-κB by impairing NF-κB suppressive-microRNA function and leads to hepatocarcinogenesis
Authors: Takata Akemi , Otsuka Motoyuki , Kudo Yotaro , Kojima Kentaro , Maeda Shin , Tateishi Keisuke , Ikenoue Tsuneo , Ijichi Hideaki , Hirata Yoshihiro , Yoshida Haruhiko , Omata Masao , Koike Kazuhiko
Published: February 2011

Hepatocellular carcinoma is the third leading cause of cancer mortality worldwide, but the molecular mechanisms in tumorigenesis remain largely unknown. Here, we show that DDX20, a DEAD-box protein identified as a tumor suppressor in a recent comprehensive in vivo screening of murine liver cancer, might have a role in human hepatocellular oncogenesis through previously undefined mechanisms. DDX20 expression was frequently decreased in hepatocellular carcinoma cells compared with the hepatocytes in the noncancerous liver tissues from the same patient, as determined by immunohistochemistry using tissue arrays. DDX20-knockdown cells showed enhanced NF-κB activity and higher interleukin-6 expression, a cytokine known to be related to hepatocarcinogenesis. Our microRNA library screening revealed that several microRNAs expressed in hepatocytes, such as miR-140, normally suppress NF-κB activities. The deficiency of DDX20, a component of microRNA-containing ribonucleoprotein complexes, impaired microRNA function and this led to the impairment of the NF-κB-suppressive microRNA function, and consequently, enhanced NF-κB activity. These results indicate that DDX20 deficiency enhances NF-κB activity by impairing NF-κB-suppressive microRNA function and may contribute to hepatocarcinogenesis. “Functional impairment of microRNAs” identified here, induced by aberrant expression or functional abnormalities of the molecules involved in the microRNA pathway, may also be one of the mechanisms of oncogenesis even in other organs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2065.
Abstract 2069: Combinatorial microRNA activity associated with breast cancer metastasis is revealed through computational analysis of target genes and pathways
Authors: Dombkowski Alan A. , Sultana Zakia , Jamil Hasan , Craig Douglas
Published: February 2011

A recent report identified six microRNAs with repressed expression in highly metastatic MDA-MB-231 breast cancer cell derivatives (miRs 335, 126, 206, 122a, 199a*, and 489). Expression of all six miRs was consistently decreased in cells that produced aggressive lung and bone metastases, as compared to parental cells, indicating that metastatic colonization is exacerbated by the concurrent loss of multiple miRs from this set. With the hypothesis that target genes involved in metastatic progression are regulated by cooperative activity of these six microRNAs, we performed a computational analysis to identify transcripts having multiple binding sites targeted by the collective set of miRs. Using miR-AT!, a new web-based computational tool developed in our laboratory, we identified 767 target transcripts with multiple sites, and we characterized the biological processes and pathways associated with the target genes. Among cellular processes enriched with genes from this set are regulation of cell cycle, cytoskeleton organization, and regulation of cell adhesion. Importantly, perturbation of these processes was recently noted as a common feature among all metastatic tumors. We have identified subsets of the six miRs that are predicted to regulate the genes in each of these critical cellular processes. Additionally, we analyzed signaling pathways downstream of the collective set of target genes to identify key regulatory molecules where the networks converge. The top scoring nodes were cyclin D1 and c-myc, suggesting that dysregulation of target genes due to the collective loss of the microRNAs may have a focused effect on the activity of these two key genes. Access to miR-AT! is available at www.mir-at.org.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2069.
Abstract 2074: Association of MnSOD and microRNA-301a regulation with inhibition of pancreatic ductal adenocarcinoma
Authors: Li Yan , Li Xuanshe , Li Yong , Brown Russell E. , Schiffman Suzanne C. , Martin Robert C.
Published: February 2011

Background: Pancreatic cancer is the fourth leading cause of cancer-related death in the United States. Manganese superoxide dismutase (MnSOD) has been found to be low in human pancreatic ductal adenocarcinoma (PDAC). Overexpressing MnSOD in PDAC has decreased growth rate of cancer cells. microRNAs, short 20-25 nucleotide RNA molecules, can function as oncogenes or as tumour suppressors. It has been shown that microRNA-301a is specifically up-regulated in PDAC specimen and inhibition of microRNA-301a has a decreased growth rate of PDAC cells. Aim: This study is to determine the relationship between the level of MnSOD and microRNA-301a regulation regarding the growth inhibition of PDAC. Methods: we carried out two experiments. A total of 60 PDAC specimens, along with 20 normal pancreatic tissues were obtained for immunohistochemistry (IHC) and in situ hybridization (ISH) studies to evaluate MnSOD and microRNA-301a expression. All pathology findings for the specimens were scored by two pathologists who were both blinded to the subjects. We also established a xenograft PDAC mouse model to study the growth rate in vivo. Nude BALB/C mice, 6 weeks old, were injected subcutaneously with 5×106 human PDAC (PanC-1 cells transfected with micro-301a antisence and control). Tumor-bearing animals were maintained for 8 weeks. Tumor growth was followed and tumor volume (TV) was determined using the standard formula, TV = (L xW2)/2. Results: There were significant decreases of MnSOD expression in PDAC specimens (1.5±0.06) compared to normal controls (2.5±0.07; p < 0.05). microRNA-301a expression was significantly increased in PDAC specimens (0.85±0.046) compared to normal controls (0.1±0.05; p < 0.05). The animal study findings indicated that microRNA-301a antisence transfection can significantly decrease the growth rate of inoculated PDAC cells. The decrease the growth rate is associated with the increased MnSOD levels (Fig 1) and decreased NF-kappaB expression. Conclusions: 1) MnSOD levels are negatively associated microRNA-301a expression in PDAC tissues; 2) increased levels of MnSOD are associated with microRNA-301a down-regulation and inhibition of PDAC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2074.
Abstract 2076: Mammary stem cell-like miRNA expression and relation to cancer
Authors: Aydogdu Eylem , Helguero Luisa , Haldosen Lars-Arne , Williams Cecilia M.
Published: February 2011

Mammary stem cells are suggested to be the origin of breast cancer, but are elusive and vaguely characterized. Cancer may be sustained by cancer stem cells, that are resistant to common treatments like irradiation and chemotherapy.

To determine a possible association between mammary stem cells and breast cancer, we have previously made a detailed characterization of the transcriptome in murine mammary epithelial stem-like cell line (HC11) and made a thorough investigation of global gene-expression changes during stepwise differentiation using dual-color comparative microarray technique. Subsequently, we performed a cross-species comparison to reveal conserved gene expression between stem cells and subtype-specific and prognosis gene signatures. Our analysis provided a unique map of the transcriptomic changes and a number of novel mammary stem cell markers. Our comparisons showed a relationship between breast cancers of poor prognosis and stem-like gene expression. In particular the specific subtypes of basal like tumors and Luminal C types, which both have a poor prognosis, showed stem cell like gene expression pattern. We are subsequently investigating regulations of miRNAs during differentiation. We have found very strong miRNA regulation during differentation of the stem cell like cells, and are investigating the functions of key regulated miRNAs. We imply the same miRNAs could be affecting poor prognosis breast cancers.

By defining the association between mammary stem cell like cells and stem cell like tumors, we can start to understand the pathways that will need to be targeted for successful treatment of poor prognosis breast cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2076.
Abstract 2078: Deregulation of miR-92a expression is implicated in hepatocellular carcinoma development
Authors: Tanaka Masami , Tsuchida Akihiko , Shigoka Masatoshi , Aoki Tetsuya , Takashina Tomoki , Takanashi Masakatsu , Murakami Yoshiki , Kuroda Masahiko
Published: February 2011

MicroRNAs (miRNAs) belong to a class of the endogenously expressed non-coding small RNAs which primarily function as gene regulators. Growing evidence suggests that microRNAs have a significant role in tumor development and may constitute robust biomarkers for cancer diagnosis and prognosis. Especially, the miR-17-92 cluster is markedly overexpressed in several cancers, and is associated with the cancer development and progression. In this study, we have demonstrated that miR-92a is highly expressed in hepatocellular carcinoma (HCC). In addition, the proliferation of a HCC cell line HepG2 was inhibited by the anti-miR-92a antagomir. On the other hand, we have found that the relative amount of miR-92a in the plasmas from HCC patients is decreased compared with that from the healthy donors. Interestingly, the amount of miR-92a was elevated after surgical treatment. Thus, although the physiological significance of the decrease of miR-92a in plasma is still unknown, deregulation of miR-92 expression in cells and plasma should be implicated in the development of HCC.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2078.
Abstract 208: Low-dose actinomycin D decreases Mcl-1 expression and acts synergistically with ABT-737 to induce apoptosis of SCLC cell lines
Authors: Xu Haishan , Humphries Malika , Krystal Geoffrey W.
Published: February 2011

SCLC is initially a highly chemotherapy-responsive disease but relapse and progressive development of chemotherapy resistance is the rule. Overexpression of pro-survival members of the Bcl-2 family, including Bcl-2, Bcl-XL and Mcl-1 are thought to play a role in the pathogenesis and chemotherapy resistance of SCLC. ABT-737, a BH3-mimetic drug which blocks the function of Bcl-2 and Bcl-XL, has shown single agent activity in preclinical models of SCLC and enhances the activity of a variety of cytotoxic agents. Low levels of Mcl-1 expression and high levels of Noxa expression have been characterized as markers of sensitivity to ABT-737 in SCLC and other malignancies. We have also noted that Noxa-mediated downregulation of Mcl-1 expression is absent in 4/4 SCLC cell lines examined, suggesting roles for this defect in the pathogenesis of SCLC and in resistance to ABT-737. Because of the short half-life of Mcl-1 mRNA and protein, we studied the effects of combining ABT-737 and actinomycin D, a general transcriptional inhibitor used in the treatment of Wilms tumor, sarcomas, germ cell and trophoblastic tumors. Actinomycin D decreased Mcl-1 expression in a dose-dependent fashion in the low ng/ml range, 3 logs lower than that required for general transcriptional inhibition. While exposure to actinomycin resulted in a cell line-dependent increase in Noxa expression, Noxa was not required for the decrease in Mcl-1 expression, as it occurred in the H209 cell line, which does not express Noxa, as well as the H69, WBA and H526 cell lines which do. Concentrations of actinomycin D from 0.4-4 ng/ml showed single agent activity across a panel of SCLC cell lines in 72 hr MTT viability assays and specifically induced a high grade G2-M cell cycle blockade. When combined with low micromolar doses of ABT-737, near complete loss of viability was seen with combination indices in the 0.5 range, indicating synergy by Chou-Talalay analysis, without a significant effect on MRC-5 pulmonary fibroblasts. In addition to loss of Mcl-1 expression and enhanced Noxa expression, correlates of apoptosis induced by the combination included marked Bid cleavage. Exposure to 4 ng/ml actinomycin was only required for the first 24 hrs of the combined incubation but the presence of ABT-737 was required for the full 72 hrs to see maximum loss of viability and clonogenic potential. This actinomycin exposure mimics a clinically achievable AUC of 100 ng.hr/ml after bolus administration, suggesting the feasibility of combination studies utilizing bolus actinomycin D administration.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 208.
Abstract 2082: The microRNA-101 regulates the expression of EP4 receptor in colon cancers
Authors: Chandramouli Anupama , Stankova Lenka , Davenport Jennifer M. , Shanas Renee , Owens Jenae M. , LaFleur Bonnie J. , Bhattacharyya Achyut K. , Nelson Mark A.
Published: February 2011

Colorectal cancer (CRC) is one of the most devastating cancers in the United States and is characterized by the accumulation of prostaglandin PGE2. PGE2 exerts its biological function via four transmembrane G protein coupled receptors (EP1-4), among which the prostanoid EP4 receptor is the most relevant. However, its expression in human CRC specimens has not been fully understood. Therefore, we wanted to clarify the expression of EP4 receptor in human tissue specimens. We collected a total of 40 CRC tissue specimens representing different stages of disease and performed immunohistochemistry (IHC) analysis. The specimens were graded for intensity and extent of staining of the EP4 receptor. Here we show that cancer tissues (n = 15) expressed significantly higher EP4 receptor staining as compared to normal colonic (n = 16) and adenoma (n = 9) tissues. We also performed western blot analysis on paired cancer and normal tissue specimens from this cohort and confirmed that the EP4 protein levels were elevated in cancer as compared to normal tissues whereas there was no difference in the expression of EP2 receptor. By contrast, the mRNA expression of the EP4 receptor gene (PTGER4) was down-regulated in these tumors as compared to paired normal tissue. We postulated that the increased EP4 receptor protein levels observed in tumors occurs by post transcriptional mechanisms. Therefore, to address this issue, we performed in silico analysis of the 3′UTR of PTGER4 gene for microRNA binding sites. We identified a consensus microRNA-101 (miR-101) seed sequence within the 3′UTR, proximal to the coding region, of the PTGER4 gene. Interestingly we observed a correlation between loss of miR-101 levels and increase in EP4 receptor protein levels in human CRC tissue specimens as compared to paired normal colonic tissue, by RT-PCR analysis. In order to directly assess this correlation, we ectopically expressed mir-101 plasmid in colon cancer cells and demonstrated that it could suppress endogenous levels of EP4 receptor protein. Together, these findings show that EP4 receptor is significantly elevated in human CRC and suggest, for the first time, that miR-101 may negatively regulate the expression of EP4 receptor by post-transcriptional mechanisms.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2082.
Abstract 2085: Mir-30 acts as an anti-metastatic microRNA and is downregulated in non-small cell lung cancer (NSCLC)
Authors: Regalla Kumarswamy , Mudduluru Giridhar , Ceppi Paolo , Kozlowski Miroslaw , Nikliński Jacek , Allgayer Heike
Published: February 2011

MicroRNAs (miRNAs) are small (around 22 nt) endogenous non-coding RNAs which regulate gene expression by base-pairing to the 3′-UTR of the target mRNA, causing translational repression or mRNA degradation. Recently, miRNAs have been shown to regulate cancer metastasis by virtue of their ability to co-ordinately repress multiple target genes. Epithelial-mesenchymal transition (EMT), which is characterized by repression of E-cadherin expression, and increased cell mobility, plays a very important role in cancer metastasis and fibrosis development. Recent evidence suggests that microRNA-30 (miR-30) acts as an antifibrotic agent and reduces tumor weight in animal models. In the present study, we have identified that the miR-30 family of miRNAs may function as anti-metastatic miRNAs by inhibiting EMT. We have studied the expression of miR-30 in various NSCLC cell lines and found that miR-30 expression is inversely proportional to the invasive capabilities of the cells. Correspondingly, all ‘mir-30 high’ cells are E-cadherin (epithelial marker) positive and N-cadherin (mesenchymal marker) negative, and vice-versa. ‘miR-30 low’ cell lines displayed several fold higher migratory and invasive abilities when compared with ‘miR-30 high’ cell lines. Similarly, in a chicken embryonic metastasis assay, we found that the number of metastasizing cells to lungs and livers of developing chicken was significantly reduced when miR-30 was over-expressed in ‘miR-30 low’ cells compared with mock-transfected cells. Over-expression of miR-30 also inhibited TGF-beta induced EMT in ‘miR-30 low’ cells, confirming the role of miR-30 in EMT inhibition. Search for miR-30 targets, that can modulate E-cadherin expression, using web based target analysis tools, have identified that Snail, which induces EMT primarily via transcriptional repression of E-cadherin, is a putative target of miR-30. Luciferase and Western blotting experiments have confirmed that miR-30 targets Snail. Finally, we have screened the expression of miR-30 in 47 consecutive NSCLC patients and found that miR-30 is down-regulated in tumours of about 80% of patients compared with their corresponding normal tissue. Collectively, these observations suggest that members of miR-30 are down regulated in non-small cell lung cancer, and inhibit EMT by targeting Snail.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2085.
Abstract 2086: Regulation of Axl receptor tyrosine kinase expression, invasion and metastasis by miR-34a and mir-199a/b
Authors: Mudduluru Giridhar , Ceppi Paolo , Regalla Kumarswamy , Allgayer Heike
Published: February 2011

Axl is a receptor tyrosine kinase over-expressed in different human cancers which induces proliferation, migration and invasion. In this study, we show that specific microRNAs (miR) can target the 3’-UTR of Axl and can modulate its expression. By a bioinformatics approach, we found conserved target sites for miR-34 and miR-199 within the Axl-3’-UTR at 31-49 nt and 29-56 nt, respectively. Luciferase-reporter assays with wild-type and deleted miR-34 and −199 seed sequences of Axl-3′UTR confirmed the specificity of targeting. An inverse correlation between Axl protein and miR-34a expression in a panel of non-small cell lung cancer (NSCLC), colorectal (CRC) and breast cancer (BRC) cell lines was observed, while no miR-199a or 199b expression was detected. Transient transfection of antagonist-miR or pre-miR for miR-34a and 199a significantly induced and reduced Axl-protein levels, respectively. Pre-miR transfection was able to inhibit in vitro migration and invasion of H1299 and RKO cells and, in vivo, to reduce the number of distant lung- or liver-metastasis in a chicken-embryo-metastasis assay. Moreover, methylation specific PCR on bisulfite-converted DNA obtained from the cell lines showed that the miR-34a promoter methylation status was significantly correlated with Axl expression, and that miR-199a/b promoter regions were methylated in all the cell lines tested. These results suggest that Axl receptor, together with its role in cancer cell growth, invasion and metastasis formation, can be regulated by miR34a and 199a/199b, and that one of the reasons for its over-expression in tumors can be the methylation of DNA that encodes these specific miRs which target Axl expression.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2086.
Abstract 2087: miR-483-3p is an oncogene involved in nephroblastoma and in adult tumors with activated β-catenin
Authors: Veronese Angelo , Consiglio Jessica , Visone Rosa , Zanesi Nicola , Ferracin Manuela , Lanza Giovanni , Querzoli Patrizia , Angioni Adriano , Croce Carlo , Negrini Massimo
Published: February 2011

The hsa-mir-483 locus is located within intron 2 of the IGF2 locus. Here, we show that the mature microRNA (miRNA) miR-483-3p is over-expressed and co-regulated with IGF2 in 100% of Wilms’ tumors. Additionally, several types of common adult cancers exhibit high or even extremely high levels of miR-483-3p. We show that the mutation status of the oncoprotein β-catenin is correlated with the expression of the miR-483-3p in hepatocarcinoma and colorectal cancer. These findings suggested an autonomous oncogenic function for this miRNA. Indeed, anti-microRNA oligonucleotide (AMO) against miR-483-3p could inhibit tumorigenicity of HepG2 cells. At the same time, no anti-tumor effect was elicited by inhibition of IGF2. The oncogenic properties of miR-483-3p were further supported by the finding that its ectopic expression could protect cells from apoptosis through the regulation of one important pro-apoptotic protein, Puma. Our results indicate that miR-483-3p can function as an anti-apoptotic oncogene in various human cancers and reveal a new potentially important target for anti-cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2087.
Abstract 2088: miR-133a as a tumor suppressive microRNA by targeting multiple oncogenes in head and neck squamous cell carcinoma (HNSCC)
Authors: Hanazawa Toyoyuki , Kikkawa Naoko , Nohata Nijoro , Suzuki Homare , Chiyomaru Takeshi , Enokida Hideki , Nakagawa Masayuki , Okamoto Yoshitaka , Seki Naohiko
Published: February 2011

Head and neck squamous cell carcinomas (HNSCC) represent 6% of all cancers. Despite of considerable advances in surgery, radiotherapy and chemotherapy, the overall 5-year survival rate for patients is less than 40% in advanced stage. The understanding of the new molecular pathways of HNSCC carcinogenesis would be helpful in improving diagnosis and therapy of the disease. MicroRNA(miRNA) are endogenous short non-coding RNA molecules, which regulate gene expression by repressing translation or cleaving RNA transcripts in a sequence-specific manner. miRNA over-expression could result in down-regulation of tumor suppressor genes, whereas their under-expression could lead to oncogenes activation. Expression levels of 665 human mature miRNAs were screened using the stem-loop real-time quantitative PCR method. We identified several down-regulated miRNAs based on miRNA expression signatures in HNSCC. A comparison of miRNA signatures from HSCC and our previous reports (Ichimi et al., Int J Cancer 2009, 125:342-352; Kano et al., AACR 2009, #552; Kikkawa et al., AACR 2009, #587) identified miR-133a that is down-regulated in common, suggesting that miR-133a is candidate tumor suppressor. Gain-of-function analysis revealed that miR-133a transfectant inhibits cell proliferation and cell invasion in cancer cells. For target genes silenced by miR-133a in cancer cells, we performed genome-wide screening (oligo microarray analysis and public database searche). Among the genome-wide analysis, several candidate genes were identified, such as, fascin homolog 1 (FSCN1) and glutathione S-transferase P1 (GSTP1). We focused on FSCN1, an actin-bundling protein, which induces parallel actin bundles in cell protrusions and increases cell motility in several human cancers. The miR-133a which has conserved sequence in the 3′UTR of FSCN1, inhibited of FSCN1 expression. The signal from a luciferase reporter assay was significantly decreased at miR-133a site, suggesting miR-133a directly regulate FSCN1. An FSCN1 loss-of-function assay found significant cell growth and invasion inhibition in HNSCC cells implying an FSCN1 is associated with HSCC carcinogenesis. Tumor suppressive miRNAs and target oncogenes may provide new insights into the mechanisms in cancer. Our findings have therapeutic implications and may be exploited for future HSCC treatments.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2088.
Abstract 2090: MicroRNAs 221/222 and genistein mediated regulation of ARHI tumor suppressor gene in prostate cancer
Authors: Chen Yi , Deng Guoren , Majid Shahana , Saini Shranjot , Zaman Saif , Khatri Gaurav , Suh Seong O. , Liu Jan , Tanaka Yuichiro , Dahiya Rajvir
Published: February 2011

ARHI, an imprinted tumor suppressor gene, is expressed in normal immortalized prostate epithelial cells, but is dramatically down-regulated in prostate cancer cell lines. Here we investigated the mechanisms of ARHI silencing in prostate cancer through microRNA and genistein (a dietary isoflavone from soy) mediated pathways. First, we evaluated ARHI mRNA levels by real time PCR using Q-PCR slice (48 prostate tumor and normal adjacent tissues) and protein levels by immunostaining of prostate tissue array (36 cancer, 8 matched normal adjacent tissue and 4 bone metastases). Both experiments showed down-regulation of ARHI in prostate cancer tissues compared to adjacent normal tissues. To understand the functional significance of ARHI down-regulation with regard to tumorigenesis, ARHI was over-expressed in prostate cancer PC-3 cells by introduction of ARHI plasmid followed by functional studies. Over-expression of ARHI can inhibit cell proliferation, colony formation, invasion and induced caspase-independent apoptosis. Further studies on new mechanism of ARHI down regulation showed a significant inverse relationship between ARHI and miR-221 & 222 which were up-regulated in cancer cell lines. Transfection of miR-221 & 222 inhibitors into PC-3 cells caused a significant induction of ARHI expression. A direct interaction of miR-221 or 222 with a target site on the 3′UTR of ARHI was confirmed by a dual luciferase pMIR-REPORT assay. Finally, we also found that genistein up-regulated ARHI by down regulating miR-221 & 222 in PC-3 cells. In conclusion, ARHI is a tumor suppressor gene down regulated in prostate cancer, and over-expression of ARHI can inhibit cell proliferation, colony formation and invasion. This study demonstrates for the first time that prostate cancer cells have elevated levels of miR-221 & 222 which bind to the 3′UTR of ARHI contributing to its down regulation. Genistein, a potential non-toxic chemopreventive agent restores expression of ARHI and may be an important dietary therapeutic agent for treating prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2090.