bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4037: Higher Expressions of miR-21, miR-106a, miR-181b, and miR-203 are Associated with Poor Prognosis in Colorectal Cancer Patients
Authors: Bovell Liselle , Katkoori Venkat R. , Shanmugam Chandrakumar , Lee Choo H. , Meleth Sreelatha , Bumpers Harvey L. , Grizzle William E. , Manne Upender
Published: February 2011

Background: MicroRNAs (miRNAs) are a family of small (20- to 24-nucleotide) non-coding RNAs that regulate gene expression at the post-transcriptional level. Although miRNAs are potential prognostic biomarkers in colorectal cancers (CRCs), their value based on the patient race/ethnicity has not been examined. Thus, this study evaluated the prognostic value of miRNAs for CRCs in African-Americans (Blacks) and non-Hispanic Caucasians (Whites).

Methods: TaqMan® microRNA assays and qRT-PCR were used to quantify the expression profiles of a panel of miRNAs (miR-20a, miR-21, miR-106a, miR-181b, and miR-203) in CRCs and corresponding benign tissues collected from 104 Blacks and 114 Whites. The expression profiles were correlated with patient overall survival, and p values were calculated by the log-rank test.

Results: For both racial groups, all five miRNAs had higher expression in CRCs (>1.0 fold; range, 1.23 to 7.52) compared to their corresponding controls. Univariate Kaplan-Meier analyses demonstrated that increased expression of miR-106a was associated with decreased overall survival of both Blacks and Whites (p=0.040 and p=0.049, respectively). Increased expressions of miR-181b and miR-203 were each significantly associated with short patient survival of Blacks (p=0.009 and 0.003, respectively), but not Whites (p=0.117 and p=0.071, respectively). When Blacks and Whites were combined, over-expression of miR-21 was associated with poor patient survival (p=0.049), but statistical significance was not reached when the values were analyzed based on race. In contrast, increased expression of miR-20a was not associated with survival of either Blacks (p=0.193), Whites (p=0.242), or the combined patient population (p=0.072).

Conclusions: These findings suggest that, for all CRC patients, over-expression of miR-21 and miR-106a are associated with poor prognosis, whereas over-expression of miR-181b and miR-203 are associated with poor outcomes for Blacks. These findings suggest that, in prognostic assessment of miRNAs in CRC, patient race/ethnicity should be considered. These studies were supported by NCI/NIH grants, U54-CA118948 and R01-CA098932-01.

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 4037.
Abstract 4042: MiR-200c sensitizes colorectal cancer cells to 5-FU through Bcl-2-involved apoptotic pathway
Authors: Qian Xuemin , Howell Paul , Li Xiaobo , Qin Xin , Gao Lin , Xi Yaguang
Published: February 2011

Effective clinical management of colorectal cancer is still a major challenge in cancer care. MicroRNA is a set of newly-discovered, non-coding small RNA molecules that significantly contribute to a number of critical biological events including cell proliferation, apoptosis development, as well as tumorigenesis. Our group has previously identified a number of microRNA candidates associated with potential prognostic value in colorectal cancer patients. In this study, we investigated miR-200c and validated its impact on 5-FU chemosensitivity in colorectal cancer. Human colorectal cancer cell lines HCT116 and HCT15 were employed in this study. Pre-miR-200c precursor molecules and negative control (Ambion, Inc.) were transiently transfected into these cell lines and cell viability was examined using WST-1 assay (Roche, Inc.) after 5-FU treatment for 24 hours. The results indicate that miR-200c can potentially sensitize colorectal cancer cells to 5-FU up to 25% compared to controls. Also, cells over-expressing miR-200c have an approximately 50% higher apoptotic rate than controls based on flow cytometry results. Bioinformatics predicts Bcl-2 is a putative mRNA target of miR200c. Bcl-2 is an integral membrane protein located mainly on the outer membrane of mitochondria. Overexpression of Bcl-2 has been demonstrated to prevent cells from undergoing apoptosis in response to a variety of stimuli. In this study, we further experimentally validated the repressive abilities of miR-200c on Bcl-2 expression using Western Blot and Luciferase reporter assay. Our results demonstrates that miR-200c is a potential biomarker for predicting chemotherapeutic response in colorectal cancer and functions by repressing Bcl-2 expression and inducing apoptotic susceptibility to 5-FU. These results hold great promise for moving miRNAs toward future clinical application and contribute to the ultimate goal of developing or improving therapeutic intervention for patients with colorectal 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 4042.
Abstract 4044: microRNA mimics as cancer therapeutics
Authors: Daige Chritopher L. , Fields Robert B. , Li ChuanYu , Cantley Will , Akinc Akin , Bhat Balkrishen , Marcusson Eric G. , Linsley Peter S.
Published: February 2011

microRNAs are small non-coding RNAs that regulate gene expression post-transcriptionally. The mRNA targets of microRNAs are usually determined based on complementarily to the 5′ end of the microRNA; the area termed the seed sequence. One microRNA can have more than 100 mRNA targets that are usually inhibited by ∼50%. The expression of microRNAs has been shown to be dysregulated in numerous cancer types. These dysregulated microRNAs can act to either positively or negatively regulate initiation, proliferation and metastasis of cancer cells. Some of the most important pathways in cancer biology (e.g. the Ras and p53 pathways) have been shown to interact with microRNAs. Therefore, both inhibiting and replacing microRNAs have therapeutic potential in oncology. We are using technology similar to that used for antisense oligonucleotides and lipid nanoparticle formulated siRNAs, both of which are currently in clinical trials for cancer, to determine the best potential microRNA based therapeutics to develop as drugs. In this report we will focus on our attempts to introduce mimics of microRNAs known to be down-regulated in tumors (e.g. miR-34a). Initial experiments were performed in wild-type mice and demonstrated the ability to deliver microRNA mimics to the liver and to affect gene expression in a seed sequence-dependent manner. These results, along with effects of successful microRNA introduction into an orthotopic xenograft model of hepatocellular carcinoma will be presented.

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 4044.
Abstract 4054: Prognostic value of microRNAs expression in diffuse large B cell lymphoma
Authors: Kushnir Michal , Lahav Meir , Ram Ron , Gilad Shlomit , Zepeniuk Meirav , Cohen Lahav , Ezagouri Meital , Baery Osnat , Spilberg Ofer , Feinmesser Meora , Aharonov Ranit , Yanai Gila Lithwick
Published: February 2011

Background: Diffuse large cell lymphoma (DLBCL) is heterogeneous in its clinical sequel. Approximately 50% of patients relapse after treatment and many succumb to the disease. Risk stratification of these patients is based mainly on the International Prognostic Index (IPI), while molecular profiling is not widely used for prognostication. Thus, there exists a need for identification of additional biomarkers that can be used as prognostic indicators for patients with DLBCL. Here we propose microRNAs, a family of small non-coding regulatory RNAs involved in cancer development, as an emerging class of potential biomarkers for risk stratification of DLBCL.

Methods: Expression profiling of microRNAs was performed on RNA extracted from biopsy specimens of 83 DLBCL patients diagnosed and treated at Rabin Medical Center, using custom microRNAs microarrays. All patients were treated with CHOP or CHOP like therapy and were dichotomized into a “good prognosis” group (patients that achieved complete remission and no relapse within 5 years) and a “bad prognosis” group (patients that were either resistant to treatment or relapsed within 9 months). Expression levels were compared between the two groups of samples and statistically analyzed. The expression of differential microRNAs was verified using a microRNA qRT-PCR platform.

Results: We found that several microRNAs are significantly differentially expressed in tumors from patients with bad prognosis compared with patients with good prognosis. Proprietary microRNA qRT-PCR showed a high correlation to microarrays and similar separation into prognosis groups based on miR expression.

Conclusion: In summary, according to the results of this study, miRNA expression can serve as a novel tool for risk stratification and prognosis prediction of DLBCL patients. This approach can potentially be used to tailor treatment and further management to specific patient needs.

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 4054.
Abstract 4055: Real-time quantitation of primary transcripts of microRNA genes
Authors: Liang Yu , Hu Fangqi , Langit Emanuel , Lu Julia , Brzoska Pius , Chen Caifu
Published: February 2011

Regulation of mature microRNA (miRNA) expression has been shown during the course of biogenesis under different contexts: from transcription, processing of pri-miRNA to pre-miRNA by Drosha, to processing of pre-miRNA to mature miRNA by Dicer. A mature miRNA can be processed from two or more stem-loop precursor loci, which are denoted with numeric suffixes such as −1, −2, etc. As of the 14th Release of the miRBase, there are 154 genes among the 750 miRNA genes that belong to this category. To interrogate regulation of miRNA expression at the transcriptional level for those that can be processed from multiple genomic locations, it requires assays to specifically quantitate the precise locus of interest. We have developed a pipeline to design and select real-time RT-PCR assays to quantitate primary transcripts of miRNA genes. These assays are designed against genomic DNA sequence so the genomic DNA needs to be removed by prior enzyme treatment. We showed that the TaqMan® Pri-miRNA Assays have minimally detectable or no background using either cell lysates or commercial tissue total RNA. Excellent linearity was seen using cDNA synthesized from total RNA, or using 1ng/ul or greater of genomic DNA. The specificity of the assays to distinguish loci that are processed into identical mature miRNA was demonstrated using subcloned plasmids, as well as using breast cancer cell lines that showed concordant expression patterns among such loci with published data in human breast cancer. The TaqMan® Pri-miRNA Assays will be a useful tool to identify the precise loci that contribute to the mature miRNA expression of interest at the transcription level, so as to further investigate the regulatory mechanisms for the differential 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 4055.
Abstract 4060: Radiation-inducible silencing of uPA and uPAR in vitro and in vivo in meningioma
Authors: Gogineni Venkateswara Rao , Nalla Arun Kumar , Gupta Reshu , Gorantla Bharathi , Gujrati Meena , Dinh Dzung H. , Fassett Daniel , Klopfenstein Jeffrey D. , Rao Jasti S.
Published: February 2011

Stereospecific radiation treatment offers a distinct opportunity for temporal and spatial regulation of gene expression at tumor sites by means of inducible promoters. To achieve this, a plasmid, pCArG-U2, was constructed by incorporating nine CArG elements (in tandem) of EGR1 gene upstream to uPA and uPAR siRNA oligonucleotides in a pCi-Neo vector. Radiation-induced siRNA expression was detected in a meningioma cell line (IOMM-Lee). Immunoblotting and RT-PCR analyses confirmed downregulation of uPA and uPAR. A similar effect was observed in transfected cells followed by H2O2 treatment. Moreover, pre-treatment of transfected cells with N-Acetyl L-Cysteine blocked the silencing of uPA and uPAR, which further confirmed the oxidative damage-mediated downregulation. Cell proliferation assays and western blot analysis for apoptotic molecules confirmed cell death in a radiation-inducible fashion. Migration and Matrigel invasion assays also revealed a marked decrease in the migration and invasive behavior. Immunocytochemistry showed a marked decrease in uPA and uPAR levels in the transfected and irradiated cells. H&E staining revealed a decrease in the pre-established tumor volume among the animals treated with pCArG-U2 and radiation. Immunohistochemistry of the brain sections established with intracranial tumors also revealed a marked decrease in uPA and uPAR in a radiation-inducible fashion. Altogether, our data suggest pCArG-U2 as a suitable candidate for radiation-inducible gene 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 4060.
Abstract 4065: Suppression of mannosylphospho dolichol synthase (DPMS) expression by vector-based siRNA technology reduces angiogenesis and inhibits tumor growth
Authors: Zhang Zhenbo , Banerjee Aditi , Santiago Jesus , Baksi Krishna , Banerjee Dipak K.
Published: February 2011

Suppression of mannosylphospho dolichol synthase (DPMS) expression by vector-based siRNA technology reduces angiogenesis and inhibits tumor growth

Zhenbo Zhang1, Aditi Banerjee1, Jesus Santiago1, Krishna Baksi2 and Dipak K. Banerjee1. 1Dept Biochem, Sch Med, Univ Puerto Rico, San Juan, PR and Dept Anatomy and Cell Biol, Sch Med, Universidad Central del Caribe, Bayamon, PR, USA.

Mannosylphospho dolichol synthase (DPMS), a 31kDa phosphoprotein is essential for the synthesis of lipid-linked oligosaccharide (LLO; Glc3Man9GlcNAc2-PP-Dol), a pre-requisite for asparagine-linked (N-linked) protein glycosylation. Earlier observations from our laboratory have suggested that up-regulation of DPMS activity plays an important role in capillary endothelial cell proliferation and consequently in angiogenesis. Our recent study suggests that blockade of N-linked glycosylation by tunicamycin may provide a novel therapeutic approach for the treatment of cancers. However, the roles of suppressing DPMS expression by genetic manipulation and its impact on angiogenesis and tumor cell proliferation have not been investigated. We report here that knocking down DPMS expression by vector-based short interfering RNA technique dramatically reduced the N-linked glycosylation of protein in endothelial cells and inhibited angiogenesis and cell proliferation. After isolating DPMS overexpression and siRNA stable cell lines, DPMS expression in these cells has been confirmed by fluorescence microscopy, western blotting, QRT-PCR and measuring the catalytic activity. All detected elevated DPMS level in overexpressing clone but is down-regulated in cells silenced with siRNA. Fluorescence microscopy with WGA supported high expression of complex glycans on the cell surface overexpressing DPMS and is considerably reduced in cells DPMS silenced with siRNA. Two-dimension gel combined fluorescence with FITC-labeled Con A and Texas-Labeled WGA also support this observation. Overexpressing DPMS clone proliferates faster compared to the cells whose DPMS has been silenced with siRNA. This is corroborated with the cell proliferative antigens Bcl-2 and PCNA expression monitored by western blotting and/or by QRT-PCR. In wound healing assay DPMS overexpression facilitated faster cell migration than its siRNA clone. Thus, DPMS exerts influence on capillary endothelial cell proliferation and may serve as a modulator of angiogenesis. We, therefore, conclude that DPMS may serve as a potential target for developing anti-angiogenic therapeutics treating cancer. Supported by Susan G. Komen for the Cure BCTR 056206 (DKB) and NIH/NCRR/RCMI G12-RR0303 (KB) grants.

1

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 4065.
Abstract 4066: Benzyl isothiocyanate causes degradation of the aryl hydrocarbon receptor in hepa1c1c7 cells
Authors: Kioko Stephen M. , Kirlin Ward G.
Published: February 2011

The biotransformation and detoxification of many carcinogens, including 2,3,7,8-tetrachlorodibenzodioxin, benzo [a] pyrene and 3-methylchloranthrene is dependent upon the activity of phase I and II enzymes, particularly CYP1A1 and NQO1. The transcriptional activation of these enzymes is regulated by the aryl hydrocarbon receptor (AhR) and Nuclear Factor-Erythroid 2-Related Factor. These transcription factors reside in the cytoplasm and when activated, shed their respective chaperones and translocate into the nucleus where they interact with nuclear proteins involved with transcription of genes possessing xenobiotic (XRE) and antioxidant (ARE) response elements. We have previously observed that the phytochemical benzyl isothiocyanate (BITC) induces nuclear translocation of the AhR independently of traditional ligands in Hepa1c1c7 cells and also induces AhR heterodimerization with ARNT. These observations suggest that BITC might bind the AhR and affect activity of these enzymes as well as the transcriptional regulation of XRE-responsive genes. In the current study we investigated whether BITC causes degradation of the AhR. This is important because it provides insight into potential mechanisms by which BITC affects AhR transcriptional regulation and the activity of phase I and II enzymes. Hepa1c1c7 cells were treated for 4 hours with DMSO alone, 10 μM BITC, 10 μM BNF or a combination of both 10 μM BITC and 10 μM BNF. Whole cell lysates were prepared and AhR immunoprecipitated and analyzed by western blot. Results showed that AhR was rapidly degraded in cells treated with BITC alone compared to controls. Furthermore, BITC enhanced degradation of the receptor in cells treated with a combination of both BITC and BNF compared to cells treated with BNF alone. In other experiments, Hepa1 cells were treated for 16 hours with BNF alone or a combination of 1, 2, 5, 10 and 20 μM BITC. Isolated proteins analyzed by western blot indicated that BITC enhanced degradation of the AhR in a dose dependent manner in cells treated with both BITC and BNF compared to BNF-only treated cells. Cumulatively, these data show that BITC degrades the AhR, which might decrease its function in transcription of XRE responsive genes. The results also suggest that increased degradation of the AhR might affect the activity of CYP1A1 and NQO1 thus affecting biotransformation and detoxification of carcinogens. They also provide preliminary evidence into one of the potential mechanisms by which BITC is chemopreventive. (Support: NIH MBRS/GM028248; RCMI RR033032).

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 4066.
Abstract 4068: Reciprocal regulation of RNase-L and tristetraproline modulates the induction of proinflammatory cytokines
Authors: Hassel Bret A. , Li Xiao-Ling
Published: February 2011

The endoribonuclease RNase-L is the terminal component of an interferon-regulated RNA decay pathway that functions in antiviral and antiproliferative/tumor suppressor activities. We recently identified a novel role for RNase-L in the induction of tumor necrosis factor-a (TNFa). The induction of TNFa is an important mediator of inflammation, therefore the RNase-L-dependent regulation of TNFa may contribute to its tumor suppressor activity. In the current study, we sought to investigate the mechanism of this regulation. The induction of TNFa mRNA was diminished in the absence of RNase-L suggesting that RNase-L targeted a negative regulator of TNFa expression. The RNA binding protein tristetraproline (TTP) is a critical regulator of TNFa that enhances the degradation of its mRNA; thus TTP represented a viable target mRNA by which RNase-L may indirectly modulate TNFa expression. Consistent with this model, inhibition of RNase-L expression using knockdown or knockout strategies enhanced TTP induction by interferon and Toll-Like Receptor agonists and reduced levels of TTP target mRNAs (e.g. TNFa). In contrast, ectopic RNase-L expression diminished TTP expression. Importantly, the stability of TTP mRNA inversely correlated with RNase-L expression, suggesting that the TTP transcript is a direct target of RNase-L. Interestingly, analysis of RNase-L expression in lipopolysaccharide-stimulated macrophages demonstrated a downregulation of RNase-L that corresponded with TTP induction, suggesting that TTP may regulate RNase-L. Indeed, TTP overexpression decreased the half-life of RNase-L mRNA, and pharmacologic MAP kinase inhibitors that block TTP activity stabilized RNase-L mRNA. Taken together, these findings suggest that RNase-L and TTP represent reciprocally regulated components of a post-transcriptional network that functions to tightly control the expression of proinflammatory cytokines, and may influence the antitumor immune response.

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 4068.
Abstract 4075: Tolfenamic Acid Inhibits Prostate Cancer Cells Growth and Tumor Development in Orthotopic Mice
Authors: Mahammad Riyaz B. , Madero-Visbal Rafael , Baker Cheryl H. , Colon Jimmie , Safe Stephen , Abudayyeh Ala , Abdelrahim Maen
Published: February 2011

Introduction: Transcription factors, Specificity proteins (Sp) play a critical role in growth and metastasis of cancer and there is clear evidence that Sp1 expression is a negative prognostic factor for survival in some cancer patients. Sp proteins regulate the expression of several genes, including PSA gene and c-Met which are associated with prostate cancer. Nonsteroidal anti-inflammatory drug, tolfenamic acid (TA) induces the degradation of specific Sp proteins (Sp1, Sp3 and Sp4) which play critical role in tumor growth and metastasis. Sp proteins also regulate the expression of survivin, a member of inhibitor of apoptosis gene family which is associated with resistance to chemotherapy and radiotherapy in several cancers. We have evaluated the anti-cancer activity of TA in prostate cancer.

Methods and Results: Human prostate cancer cells (PC-3, DU-145 and LNcaP) were grown in the presence of DMSO (vehicle) or various concentrations (25, 50 and 100 µM) of TA and cell number and viability were measured at 24, 48, and 72 h post-treatment. Cell viability was measured using Cell Titr Glo kit (Promega) and the experiments were performed in triplicate. Cell lysates were prepared from the cells exposed to DMSO or 50 µM TA for 48 h and evaluated the expression of Sp proteins (Sp1, Sp3 and Sp4), and c-Met through Western blot analysis. For in vivo studies nude mice were injected with PC-3 Luc cells into the right prostate. Treatment (vehicle or 50 mg/kg TA/2 days) was initiated after one week and tumor growth was continuously monitored through Kodak imaging system. 4 weeks after the treatment tumors were isolated, measured and processed for immunohistochemistry to analyze the expression of Sp proteins and c-Met. In vitro studies show a significant decrease in the cell proliferation and viability and these results are dose and time dependent in all the cell lines. Furthermore, TA treatment for 48 h significantly decreased the expression of Sp1, Sp3 and c-Met in PC-3 cells. In vivo studies revealed that TA significantly decreased the tumor weight and volume which was associated with a low expression of Sp1, Sp3 and c-Met in tumor sections as evaluated by immunohistochemistry.

Conclusion: Targeting key transcription factors such as Sp proteins using a small molecule compound, TA is a new strategy for the treatment of prostate cancer and our results are very crucial in the development of a safe, cost effective drug for the treatment of this devastating disease. Studies to test the effect of TA on critical genes associated with prostate cancer and TA's impact on developing higher response to radiation therapy are underway.

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 4075.
Abstract 4078: Over-expression of FOXM1 is associated with cell migration/invasion in ovarian cancer
Authors: Lok Gabriel Tm , Chan David W. , Liu Vincent Ws , Ngan Hextan Ys
Published: February 2011

FOXM1 is a typical transcription factor regulating the S- and M-phase progression in cell cycle. It is frequently over-expressed and is associated with tumor stages in numerous human cancers. The influence of FOXM1 expression on cell proliferation has been well studied, yet its impact on other tumor characteristics in ovarian cancer, like cell migration/invasion remains unclear. In the present study, we demonstrated that FOXM1 was significantly up-regulated in ovarian cancer cell lines at both RNA and protein level by semi-quantitative RT- PCR and Western blot analyses. Immunohistochemical analysis showed that FOXM1 was highly correlated with the high-grade ovarian cancer. To examine the effect of FOXM1 on ovarian cancer cell motility, the FOXM1 specific inhibitor, thiostrepton, was used to reduce FOXM1 expression in OVCA433 cells. The FOXM1 reduced OVCA433 cells, as a result, showed a significant decrease in cell migration using wound healing assay. By using MEK specific inhibitor, U0126, we demonstrated that the inhibition of ERK activity caused a reduction of FOXM1 level in a time dependent manner and significant reduction of cell migration rate in the wound healing assay in OVCA433. In sum, FOXM1 is over-expressed and regulated by constitutive activated ERK activity in ovarian cancer cells. Our data also suggested that the ERK /FOXM1 signaling axis is crucial in regulating migration/invasion of ovarian cancer cells. Further studies on the molecular mechanism of FOXM1 in the regulation of ovarian cancer cell migration/invasion are warranted.

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 4078.
Abstract 4091: Epigenetic silencing of microRNA in the pathobiology of CLL
Authors: Sampath Deepa , Liu Chaomei , Wierda William G. , Keating Michael J.
Published: February 2011

MicroRNAs (miRs, miRNA) are a class of non-coding RNA that control gene expression at a post-transcriptional level. Specific and aberrant miRNA signatures distinguish indolent and relapsed forms of chronic lymphocytic leukemia (CLL). In this study, we focused on the expression and cellular targets of a set of microRNA (miR15a, miR-16, miR-29b and miR-101) that are differentially expressed in indolent and refractory disease. The tumor suppressor microRNAs, miR-15a and miR-16, are downregulated in 68% of all CLL and associated with an indolent course. These miRNA are located on an intron of the dleu2 gene within a genomic locus (chr13q) that is frequently deleted in leukemia. However, deletions in 13q usually involve only one allele and do not adequately account for the low expression levels of miR-15a and miR-16. We hypothesized that aberrant recruitment of the histone deacetylases could be a novel mechanism that drives the silencing of miR-15a and miR-16 in CLL. Correspondingly, exposure to a variety of small molecule histone deacetylase inhibitors led to the re-expression of miR-15a and miR-16 in 30% of samples that undergo silencing. We then evaluated the cellular targets of miR-15a and miR-16 in CLL. Ectopic expression of miR-15 and miR-16 in primary leukemia cells led to a loss of Bcl-2 and Mcl-1. However, pharmacological re-expression of these miRNA led to decreases in Mcl-1 protein, with no loss in Bcl-2. Losses in the levels of Mcl-1 were associated with induction of cell death in these leukemia cells. In contrast to miR-15a and miR-16, the microRNAs miR-29b and miR-101 are well expressed in indolent disease but become silenced in relapsed/refractory disease by unknown mechanisms. We identified that exposure to a variety of histone deacetylase inhibitors induced the re-expression of miR-29b and miR-101 in 30-50% of the downregulated samples. Chromatin immunoprecipitation experiments demonstrated that deacetylase inhibition led to an increase in permissive chromatin marks at the gene promoters for these miRNA. miR-29b is known to target Mcl-1, Tcl-1 as well as the DNA methyltransferases 3a and 3b. Ectopic expression of miR-29b in CLL cells led to the decrease of Mcl-1 protein. In parallel, expression of miR-101 led to the inhibition of EZH2 expression. Correspondingly, pharmacological re-expression of miR-29b and miR-101 was associated with losses in Mcl-1 and EZH2 in CLL cells and apoptotic death of these leukemia cells. These results identify a role for epigenetic mechanisms in the pathological silencing of miRNA in both indolent and relapsed CLL. They also offer a novel therapeutic strategy by which to reverse epigenetic silencing of miRNA for therapeutic benefit.

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 4091.
Abstract 4095: Aberrant expression of epigenetically regulated microRNAs in liver cancer
Authors: Wong Chun-Ming , Wong Carmen Chak-Lui , Tam Aegean Hoy Kam , Ng Irene Oi-Lin
Published: February 2011

Liver cancer (hepatocellular carcinoma, HCC) is a major malignancy worldwide. Despite definite improvements in the outcome of patients with HCC, the overall prognosis of this cancer is still unsatisfactory because of late presentation and frequent tumor recurrence after surgical resection. New diagnostic and adjuvant treatment modalities are much awaited. MicroRNAs (miRNAs) are a class of small (∼22 nt) single-stranded noncoding RNAs widely expressed in eukaryotic cells. miRNAs play an important role in modulating gene expression by promoting mRNA degradation or translational repression. There is mounting evidence that miRNAs are implicated in almost all major physiological processes, including cell proliferation, differentiation, apoptosis, and human carcinogenesis. Epigenetic alterations including aberrant DNA methylation and histone modification are important in regulating chromatin structure and gene expression. In this study, we investigated the role of epigenetic alterations in miRNA deregulation in human HCC. miRNA expression changes of HCC cell lines upon treatment with DNA demethylating agent (5-Aza-dC) and histone deacetylase inhibitor (Trichostatin A, TSA) were compared with stem-loop RT-qPCR based TaqMan low density array. In total, 98 miRNAs were remarkably up-regulated upon 5-Aza-dC/TSA treatment. Among these, 35 miRNA were commonly upregulated by treatment with the epigenetic inhibitors in both cell lines. The majority of these miRNAs was either associated with promoter CpG islands or located on imprinted regions. Expression analysis revealed that these epigenetically regulated miRNAs were frequently deregulated in human HCCs, and their expression pattern was able to distinguish primary HCCs from their corresponding non-tumorous livers. Interestingly, CpG island-associated miRNAs tended to be down-regulated in human HCCs, whereas, miRNAs located on imprinted regions were frequently overexpressed in human HCC samples. Our findings suggest that aberrant epigenetic silencing and loss of imprinting may contribute to miRNA deregulation in human HCC.

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 4095.
Abstract 4103: Sphingosine kinase-1 mediates LPA-induced gastric cancer cell proliferation
Authors: Ramachandran Subramaniam , Shida Dai , Nagahashi Masayuki , Milstien Sheldon , Spiegel Sarah , Takabe Kazuaki
Published: February 2011

The lysophospholipids, lysophosphatidic acid (LPA) and sphingosine-1 phosphate (S1P) are important mediators of a variety of biological processes important for maintenance and progression of cancer. Autotaxin and sphingosine kinase 1 (SphK1), the enzymes responsible for synthesis of LPA and S1P, respectively, are elevated in many cancer types. We previously reported that LPA markedly enhanced SphK1 and S1P3 receptor mRNA levels, transactivated EGFR, and caused migration in MKN-1 gastric cancer cells [Shida et al., Cancer Res. 68:6569, 2008]. We have now shown that LPA potently stimulates proliferation of MKN-1 cells and downregulation of SphK1 or treatment with the isozyme-specific SphK1 inhibitor, SK1-I, markedly reduced LPA-mediated proliferation. The signaling pathways responsible for LPA-induced SphK1 upregulation were also investigated. U0126, a MEK1/2 inhibitor, completely abrogated LPA-induced elevation of SphK1 mRNA, while inhibitors of PI3K, protein kinase C, p38, JNK or NF-κB did not have an effect. Additionally, siRNA mediated downregulation of ERK2, but not AKT 1 or Akt2, blocked LPA-mediated SphK1 mRNA increase. These results were further confirmed by adenoviral expression of dominant negative MEK1, which inhibited LPA-induced SphK1 upregulation, while constitutively active MEK1 increased basal SphK1 mRNA levels. Furthermore, adenoviral expression of dominant negative or constitutively active AKT1 also did not change SphK1 mRNA levels. Our data implicate ERK2 as an important mediator of LPA signaling and upregulation of SphK1 and additionally, point to SphK1 and S1P production as potential therapeutic targets in gastric cancer. This work was supported by NCI grant R01CA61774 to SS and NIH grant 5K12HD055881 and Susan G. Komen for the Cure Research Foundation Career Catalyst Research Grant KG090510 to KT.

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 4103.
Abstract 4107: Human guanylate-binding protein-1 (GBP-1) in colorectal carcinoma
Authors: Britzen-Laurent Nathalie , Lipnik Karoline , Naschberger Elisabeth , Kuhn Elisabeth , Hohenadl Christine , Stürzl Michael
Published: February 2011

The human guanylate binding protein 1 (hGBP-1) is a large GTPase which mediates the anti-proliferative and anti-angiogenic effects of pro-inflammatory cytokines (IC), such as IFNγ, TNFα and IL1α/β, on endothelial cells in vitro. The expression of GBP-1 has been investigated in human colorectal carcinoma (CRC). We could show that GBP-1 is expressed in stromal endothelial cells of 32% of the CRC. This expression was correlated with a better outcome for patients with GBP-1 expression in the tumour stroma. A transcriptome analysis revealed that in the tumours, IFN-γ-induced genes including the major antiangiogenic chemokines CXCL9, CXCL10 and CXCL11 were coexpressed with GBP-1, indicating the presence of an active Th-1-like angiostatic immune reaction. However, the presence of GBP-1 was only detected in the stroma, suggesting that in tumour cells the expression of GBP-1 may be suppressed. In order to investigate the effects of GBP-1 on tumour cells, we reintroduced the protein in the DLD-1 colorectal carcinoma cell line, which does not produce GBP-1 in absence or in presence of IFN-γ. The proliferation rate was lower in GBP-1 expressing DLD-1 cells as compared to control cells. In order to better characterize the in vivo anti-tumourigenic effects of GBP-1, we developed a tumour mouse model using the DLD-1 cell line stably expressing GBP-1. Stable DLD-1 clones, which expressed GBP-1, and respective control cells (transfected with the empty vector plasmid) were xenotransplanted into immunodeficient mice. After 36 days, tumour volumes in the GBP-1 group were significantly smaller than in the control group (p<0.001). This experiment was repeated with identical outcome in two different mouse strains. Immunohistochemistry stainings and Western blot analyses of the tumours confirmed that GBP-1 was highly expressed in the GBP-1 group and absent in control tumours. These results show that GBP-1 expression can inhibit tumour growth in an animal mouse model, indicating that tumour cells may have developed mechanisms to inhibit GBP-1 expression in an IFN-γ dominated microenvironment. The responsible mechanisms are investigated in ongoing studies.

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 4107.
Abstract 4109: EphrinA2 promotes tumorigenicity through Rac1/Akt/NF-kappa B signaling pathway in liver cancer
Authors: Feng Yu-xiong , Zhao Jiang-Sha , Cheng Shu-Qun , Yuan Yunfei , Wang Xiao-fan , Xie Dong
Published: February 2011

Eph/Ephrin family, one of the largest receptor tyrosine kinase families, has been extensively studied in morphogenesis and neural development. Recently, growing attentions have been attracted on its role in initiation and progression of various cancers. However, the role of Eph/Ephrins in hepatocellular carcinoma (HCC) has been rarely investigated. In this study, we found that the expression of EphrinA2 was significantly upregulated in both established cell lines and clinical tissue samples of HCC, and the most significant increase was observed in the tumors invading into the portal veins. Forced expression of EphrinA2 in HCC cells significantly promoted in vivo tumorigenicity, while knockdown of this gene inhibited this oncogenic effect. We further found that suppression of apoptosis, rather than accelerating proliferation, was responsible for EphrinA2-enhanced tumorigenicity. In addition, EphrinA2 endowed cancer cells with resistance to TNF-alpha-induced apoptosis, thus facilitating their survival. Furthermore, we disclosed a novel EphrinA2/Rac1/Akt/NF-kB pathway contributing to the inhibitory effect on apoptosis in HCC cells. In a conclusion, our study revealed that EphrinA2 played an important role in the development and progression of HCC by promoting the survival of cancer cells, indicating its role as a potential therapeutic target in 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 4109.
Abstract 4110: Attenuation of colon cancer cell adaptation to hypoxia by cGMP-dependent protein kinase
Authors: Kwon In-Kiu , Wang Rui , Prakash Nikhil , Browning Darren D.
Published: February 2011

There is growing evidence for the antitumor effects of cGMP-dependent protein kinase (PKG) in colon cancer cells. We reported recently that ectopically expressed PKG resulted in defective angiogenesis in xenografts, and was associated with reduced levels of vascular-endothelial growth factor (VEGF). In order to better understand the therapeutic potential of this enzyme, the present work has examined the mechanism of VEGF regulation by PKG. In contrast to the SW620 xenografts, PKG had no effect on VEGF expression in SW620 cells grown under standard tissue culture conditions in vitro. However, the increase in VEGF expression observed when the cells were subjected to hypoxic stress was significantly attenuated by PKG at both the mRNA and protein levels. Using luciferase reporter assays, PKG was shown to inhibit hypoxia inducible factor (HIF) transcriptional activity in several colon cancer cell lines, including SW620, HCT116, and HT29. The attenuation of HIF by PKG reflected a more fundamental inhibition of the hypoxic response, which was supported by the ability of PKG expression to block the hypoxia-induced activation of glycolytic genes including enolase and hexokinase 2. The reduced levels of glycolytic enzymes in PKG-expressing cells relative to control cells was associated with reduced glycolysis, mitochondrial activity, and cellular ATP levels under hypoxic conditions. While the inhibition of SW620 cell proliferation occurred independently of PKG expression in a 1% O2 environment, PKG reduced cell viability and increased necrosis under these conditions. These findings demonstrate for the first time that PKG can block the adaptation of colon cancer cells to hypoxia, and highlights this enzyme for further evaluation as a potential target for colon cancer treatment.

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 4110.
Abstract 4113: Role of GP88 (Progranulin) in the proliferation and survival of non small cell lung carcinoma
Authors: Xia Guanjun , Kamimura Sayuri , Yue Binbin , Dong Jianping , Serrero Ginette
Published: February 2011

In 2009, 219,000 patients were diagnosed with lung cancer in the US and 159,000 died of the disease. It has been widely reported that results of standard treatment in lung cancer are poor except for the most localized cancers, thereby emphasizing the need to discover novel targets of lung cancer for the development of novel therapies and diagnosis. In the present study, we investigated the role of the autocrine growth and survival factor GP88.

GP88 (also known as Progranulin) is an 88kDa glycoprotein that has been shown to play a critical role in the proliferation and survival of breast cancer. Pathological studies with breast cancer cases in correlation with clinical outcome data have also shown that GP88 expression was associated in invasive ductal carcinoma with increased risk of recurrence. In the present study, we examined the role of GP88 in non small cell lung carcinoma (NSCLC). Examination of GP88 protein expression by immunohistochemistry using Oncostain 88 kit was carried out with formalin fixed paraffin embedded sections of lung cancer cases. Results showed that non small cell lung carcinoma, adenocarcinoma and squamous carcinoma stained positive for GP88 whereas small cell carcinoma sections were mostly negative. In addition, normal lung tissues did not express GP88. Based on this observation, expression and action of GP88 in NSCLC was examined using representative cell lines. We show here that GP88 stimulated proliferation and survival of A549 and H1299 cell lines. GP88 expression was upregulated in cells that were made resistant to cytotoxic drugs such as taxol and velcade. Inhibition of GP88 expression by SiRNA or GP88 action by treating the cells with AG1, an anti-GP88 monoclonal antibody resulted in an inhibition of cell migration and proliferation of these drug resistant cells and increased response to cytotoxic drugs.

Based on these results, studies were carried out to examine the effect of AG1 antibody on migration and tumor formation of A549 and H1299 cells in vitro and in vivo. Our data that will be presented here point out that GP88 is a novel therapeutic target for NSCLC.

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 4113.
Abstract 4118: Fibroblast heterogeneity in oral squamous cell carcinoma is directed by tumor epithelium and promotes keratinocyte invasion in vitro
Authors: Lim Kue Peng , Wei WenBin , Parkinson Kenneth , Prime Stephen Stewart
Published: February 2011

The transformation of human cells requires the accumulation of a broad spectrum of genetic abnormalities that typically lead to unrestrained cell growth. It has now become clear that alterations extrinsic to the tumor cell are also required and there is a considerable body of evidence to show that the cancer microenvironment plays a pivotal role in the initiation, progression and spread of cancer. In this study, we show that fibroblasts isolated from keratinocyte cultures of normal oral mucosa, epithelial dysplasia and mortal and immortal oral squamous cell carcinomas (OSCC) were heterogeneous and, using global gene expression profiling, could be grouped according to their tissue of origin. Fibroblasts from immortal OSCC, for example, showed high levels of activation and were characterized by down regulation of cell cycle-related genes and up-regulation of transmembrane-related genes. The data was validated using real time quantitative PCR to examine the expression of specific up-regulated (IGFBP7, ITGA6, p21) and down-regulated (FOXM1, MCM7) genes identified as having significant fold change in the expression analyses. Further, we examined the inter-relationship between tumor epithelial cells and tumor-associated fibroblasts. Whilst the proliferation of two target keratinocyte cell lines was stimulated by conditioned media from both normal and tumor-associated fibroblasts (in excess of keratinocyte growth in 10% serum), only fibroblasts from immortal OSCC stimulated the invasion of a non-tumorigenic keratinocyte cell line into fibroblast-rich collagen gels in organotypic cultures. Keratinocyte conditioned media stimulated the growth and activation of fibroblasts from normal oral mucosa and mortal OSCC. The results demonstrate a close inter-relationship between tumor epithelium and tumor-associated fibroblasts and show that the diversity of fibroblasts during tumor progression is not only influenced by the tumor epithelium but also, has biological significance in the promotion of tumor cell invasion in vitro.

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 4118.