bluuebunny/crossref_metadata_2025 huggingface dataset:
Further refinements of next-generation force fields — Nonempirical localization of off-centered points in molecules
Authors: Chaudret Robin , Gresh Nohad , Cisneros G. Andrés , Scemama Anthony , Piquemal Jean-Philip
Published: April 2013

In the present work, we investigate different possibilities for the nonempirical localization of nonatomic centers within the context of the design of new generation polarizable force fields. To do so, the positions of electron localization function (ELF) and electron pair localization function (EPLF) attractors and of Boys orbital centroids are determined for a set of thirteen saturated and conjugated biologically relevant molecules. We consider the similarities and differences in the representations of localized lone pairs and π delocalized systems by these approaches, as well as the effects of the basis sets and of the level of quantum chemistry (QC). All three methods give consistent results upon dealing with the localized lone pairs. Concerning aromatic systems, whereas ELF and EPLF methods give mutually consistent results, the Boys scheme breaks the symmetry by alternating the electron distributions along the C–C bonds providing a different distribution of off-centered points. We then investigate the influence of lone pair localization in the water model of the Gaussian electrostatic model/sum of interactions between fragments ab initio computed (GEM/SIBFA) polarizable force field, which embodies an explicit representation of the lone pairs. This is done, in a series of mono- and poly-hydrated Zn(II) complexes, by comparing the overlap-dependent repulsion (Erep) and charge-transfer (Ect) contributions to their QC counterparts. We resort to either the current GEM/SIBFA water lone pair internal coordinates or to the ones derived from the previously mentioned localization procedures. The latter enables closer reproductions by Erep and Ect (GEM/SIBFA) of their exchange repulsion (Eexch) and Ect QC counterparts. The present preliminary results show that QC localization procedures can be used to derive accurate, nonempirical positions for off-centered points intervening in the formulation of the overlap-dependent contributions of next-generation polarizable force fields.
Multicomponent assembly of 2-amino-4-substituted 4H-chromenes using cyclic Michael donors
Authors: Lakshmi Neelakandan Vidhya , Kiruthika Selvarangam E. , Perumal P.T.
Published: April 2013

An efficient one-pot synthesis has been disclosed to access a series of 2-amino-4-substituted 4H-chromenes from salicylaldehyde, malononitrile, and a few cyclic nucleophiles using indium trichloride as a catalyst. This methodology affords high yields of products in a shorter reaction time.
Spatial and temporal patterns in total phosphorus in south-central Ontario streams: the role of wetlands and past disturbance
Authors: O'Brien Heather D. , Eimers M. Catherine , Watmough Shaun A. , Casson Nora J.
Published: April 2013

Unraveling the cause(s) of declines in total phosphorus (TP) observed over the past three decades across many forested catchments in central Ontario remains a research priority. The objectives of this paper were firstly to test the assumption that weir water chemistry is representative of catchment changes at two adjacent headwater streams that have shown declines in TP and secondly use long-term measurements of weir water quality (1980–2008) to identify potential drivers of TP declines. Longitudinal measurements confirmed that weir water chemistry is generally representative of the entire stream channel, although channel-associated wetlands have a strong influence on TP. Additionally, processes operating in wetlands dictate seasonal patterns in TP, and differences in wetland cover explain the approximately twofold difference in average TP between the two catchments. Changes in wetland cover or processes, however, cannot explain declines in TP. Rather, declines in TP are most coherent with declines in stream potassium (K) and nitrate (NO3-N), which also occurred over the past 30 years. We suggest that coincident declines in TP, NO3-N, and K may be best explained by recovery from a disturbance event.
Baicalein Preconditioning Protects Cardiomyocytes from Ischemia-Reperfusion Injury via Mitochondrial Oxidant Signaling
Authors: Chang Wei-Tien , Li Jing , Vanden Hoek Matthew S. , Zhu Xiangdong , Li Chang-Qing , Huang Hsien-Hao , Hsu Chin-Wang , Zhong Qiang , Li Juan , Chen Sy-Jou , Vanden Hoek Terry L. , Shao Zuo-Hui
Published: April 2013

Previous studies suggest baicalein, in addition to its antioxidant effects, protects against hypoxia/reoxygenation injury via its pro-oxidant properties. We hypothesize that a brief period of baicalein treatment prior to ischemia/reperfusion (I/R) may trigger preconditioning protection via a mitochondrial pro-oxidant mechanism. Using an established chick cardiomyocyte model of I/R, cells were preconditioned with baicalein (10 μM) for 10 min followed by 10-min wash prior to I/R. Intracellular oxidants were measured using 2′, 7′-dichlorofluorescin diacetate (DCFH/DA). Cell viability was assessed by propidium iodide and apoptosis determined by DNA fragmentation. Baicalein induced a transient but significant increase of DCF fluorescence within the 10-min preconditioning period, and led to significant reduction of cell death (38.9 ± 1.8% vs. 58.7 ± 1.2% in I/R control, n = 6, p < 0.001) and DNA fragmentation after I/R. Cotreatment with N-acetylcysteine (500 μM), mitochondrial complex III electron transport chain inhibitor myxothiazol (1 μM), mitochondrial KATP channel blocker 5-hydroxydecanoate-Na (5-HD, 500 μM) or anion channel inhibitor 4′, 4′-diisothiocyanato-stilbene-2, 2′-disulfonic acid (DIDS, 200 μM) resulted in significant abrogation of oxidant increase during induction as well as the protection conferred by baicalein preconditioning. These results suggest that baicalein preconditioning exhibits significant anti-apoptotic protection against cardiomyocyte I/R injury by mitochondrial oxidant signaling, which was in part mediated by mitochondrial KATP channel and anion channel opening.
Protective Effect of Rosmarinic Acid is Through Regulation of Inflammatory Cytokine in Cadmium-Induced Ototoxicity
Authors: Kim Su-Jin , Um Jae-Young , Kim Sung-Hoon , Hong Seung-Heon
Published: April 2013

Cadmium ( Cd2+ ) is an environmental contaminant that causes a variety of adverse effects. Auditory cells are sensitive to cadmium, and the cochlea is more vulnerable to cadmium toxicity than the other parts of the auditory system. Rosmarinic acid (RA) exhibits a wide spectrum of biological activities, mainly antioxidant and anti-inflammatory activities. However, the regulatory effects of RA in the auditory system have not been elucidated. In this study, we investigated the protective effects of RA on Cd2+ -induced ototoxicity in vitro and ex vivo. The findings showed that RA inhibited Cd2+ -mediated cell toxicity, reactive oxygen species generation, interleukin (IL)-6 and IL-1β production, the translocation of the apoptosis inducing factor into the nucleus and activation of caspases-3 in an auditory cell line, HEI-OC1. In addition, RA prevented the destruction of hair cell arrays in the rat organ of Cortiprimary explants in the presence of Cd2+ . These results are expected to improve our understanding of the pharmacological mechanism of RA, and help develop potential therapeutic strategies against ototoxicity.
A CANONICAL RANK-THREE TENSOR MODEL WITH A SCALING CONSTRAINT
Authors: SASAKURA NAOKI
Published: April 2013

A rank-three tensor model in canonical formalism has recently been proposed. The model describes consistent local-time evolutions of fuzzy spaces through a set of first-class constraints which form an on-shell closed algebra with structure functions. In fact, the algebra provides an algebraically consistent discretization of the Dirac–DeWitt constraint algebra in the canonical formalism of general relativity. However, the configuration space of this model contains obvious degeneracies of representing identical fuzzy spaces. In this paper, to delete the degeneracies, another first-class constraint representing a scaling symmetry is added to propose a new canonical rank-three tensor model. A consequence is that, while classical solutions of the previous model have typically runaway or vanishing behaviors, the new model has a compact configuration space and its classical solutions asymptotically approach either fixed points or cyclic orbits in time evolution. Among others, fixed points contain configurations with group symmetries, and may represent stationary symmetric fuzzy spaces. Another consequence on the uniqueness of the local Hamiltonian constraint is also discussed, and a minimal canonical tensor model, which is unique, is given.
POINT SET DISTANCE AND ORTHOGONAL RANGE PROBLEMS WITH DEPENDENT GEOMETRIC UNCERTAINTIES
Authors: MYERS YONATAN , JOSKOWICZ LEO
Published: April 2013

Classical computational geometry algorithms handle geometric constructs whose shapes and locations are exact. However, many real-world applications require modeling and computing with geometric uncertainties, which are often coupled and mutually dependent. In this paper we address the relative position of points, point set distance problems, and orthogonal range queries in the plane in the presence of geometric uncertainty. The uncertainty can be in the locations of the points, in the query range, or both, and is possibly coupled. Point coordinates and range uncertainties are modeled with the Linear Parametric Geometric Uncertainty Model (LPGUM), a general and computationally efficient worst-case, first-order linear approximation of geometric uncertainty that supports dependence among uncertainties. We present efficient algorithms for relative points orientation, minimum and maximum pairwise distance, closest pair, diameter, and efficient algorithms for uncertain range queries: uncertain range/nominal points, nominal range/uncertain points, uncertain range/uncertain points, with independent/dependent uncertainties. In most cases, the added complexity is sub-quadratic in the number of parameters and points, with higher complexities for dependent point uncertainties.
FRACTAL ANALYSIS OF LIPASE–CATALYSED SYNTHESIS OF BUTYL BUTYRATE IN A MICROBIOREACTOR UNDER THE INFLUENCE OF NOISE
Authors: PATNAIK PRATAP R.
Published: April 2013

Microbioreactors operated in real environments are often subject to noise from the environment. This is commonly manifested as fluctuations in the flow rates of the feed streams. Previous studies with larger bioreactors have shown that noise can seriously impair the performance. Given this possibility, the effects of noise on the performance of a microbioreactor have been analyzed for the trans-esterification of vinyl butyrate by 1-butanol by immobilized lipase B to produce butyl butyrate. As in previous work for macrobioreactors, the analysis was done with (i) no noise, (ii) unfiltered noise, and (iii) noise filtered by four different methods, and the fractal dimension of the product was used as an index of the performance.

All fractal dimensions decreased with increasing dilution rates, and significant stochastic chaos was likely at low dilution rates. Of the four types of filters, the auto-associative neural filter (ANF) was the most effective in reducing chaos and restoring of smooth, nearly noise-free performance. The ANF also does not require a process model, which is a significant advantage for real systems. Simulations also revealed that even in the absence of noise, deterministic chaos is possible at low dilution rates; this underscores the importance of efficient filtering under such conditions when external noise too is present. The results thus establish the importance of noise in microbioreactor behavior and the usefulness of the fractal dimension in characterizing the effects.
PIECEWISE QUARTIC TRIGONOMETRIC POLYNOMIAL B-SPLINE CURVES WITH TWO SHAPE PARAMETERS
Authors: DUBE MRIDULA , SHARMA REENU
Published: April 2013

Analogous to the quartic B-splines curve, a piecewise quartic trigonometric polynomial B-spline curve with two shape parameters is presented in this paper. Each curve segment is generated by three consecutive control points. The given curve posses many properties of the B-spline curve. These curves are closer to the control polygon than the different other curves considered in this paper, for different values of shape parameters for each curve. With the increase of the value of shape parameters, the curve approach to the control polygon. For nonuniform and uniform knot vector the given curves have C0, G3; C1, G3; C1, G7; and C3 continuity for different choice of shape parameters. A quartic trigonometric Bézier curves are also introduced as a special case of the given trigonometric spline curves. A comparison of quartic trigonometric polynomial curve is made with different other curves. In the last, quartic trigonometric spline surfaces with two shape parameters are constructed. They have most properties of the corresponding curves.
SHARP FEATURES EXTRACTION FROM POINT CLOUDS
Authors: SLAMU WUSHOUR , CAO JUMING , YAO XINHUI
Published: April 2013

As sharp feature manipulation plays an important role in point clouds processing, a novel mean curvature flow-based framework for sharp feature extraction from point clouds is presented in this paper. That is, for input point clouds, a general purpose mean curvature flow-based point clouds smoothing operator is applied on them, thereby, obtaining a smoothing version of the original point clouds. The sharp feature points are labeled as points whose displacements between original point clouds and their smoothing version get local extreme. Implementation of our method on both synthesized and real scanned point clouds show that our methods are effective and robust for the purpose of sharp features extraction tasks.
ELECTRIC FIELD EFFECTS ON QUANTUM CORRELATIONS IN THREE COUPLED SEMICONDUCTOR QUANTUM DOTS
Authors: SHOJAEI S. , MAHDIAN M. , YOUSEFJANI R.
Published: April 2013

We study the effect of external electric bias on the quantum correlations in the array of optically excited three coupled semiconductor quantum dots. The correlations are characterized by the quantum discord and concurrence and are observed using excitonic qubits. We employ the lower bound of concurrence for thermal density matrix at different temperatures. The effect of the Förster interaction on correlations will be studied. Our theoretical model detects nonvanishing quantum discord when the electric field is on while concurrence dies, ensuring the existence of nonclassical correlations as measured by the quantum discord.
Sparse Planar Array Synthesis Using Matrix Enhancement and Matrix Pencil
Authors: Zheng Mei-yan , Chen Ke-song , Wu Hong-gang , Liu Xian-pan
Published: April 2013

The matrix enhancement and matrix pencil (MEMP) plays important roles in modern signal processing applications. In this paper, MEMP is applied to attack the problem of two-dimensional sparse array synthesis. Firstly, the desired array radiation pattern, as the original pattern for approximating, is sampled to form an enhanced matrix. After performing the singular value decomposition (SVD) and discarding the insignificant singular values according to the prior approximate error, the minimum number of elements can be obtained. Secondly, in order to obtain the eigenvalues, the generalized eigen-decomposition is employed on the approximate matrix, which is the optimal low-rank approximation of the enhanced matrix corresponding to sparse planar array, and then the ESPRIT algorithm is utilized to pair the eigenvalues related to each dimension of the planar array. Finally, element positions and excitations of the sparse planar array are calculated according to the correct pairing of eigenvalues. Simulation results are presented to illustrate the effectiveness of the proposed approach.
Analysis for Resolution of Bistatic SAR Configuration with Geosynchronous Transmitter and UAV Receiver
Authors: Jiang Yicheng , Wang Zhuoqun
Published: April 2013

Bistatical SAR with geosynchronous illuminator and unmanned aerial vehicle receiver (GEO-UAV BiSAR) has significant potential advantages in the field of continuous local observation under a dangerous environment within nearly 24 h. Due to the extreme platform velocity differences, the ellipse orbital movement of GEOSAR makes this BiSAR configuration not like the conventional spaceborne BiSAR. In this paper, based on the orbital kinetic characteristic of GEOSAR, we theoretically analyze the variations of bistatic configuration effect on common azimuth coverage and coherent accumulated time. In addition, two-dimension the resolution is deduced by geometrical configuration on the basis of gradient method. The simulations show that the appropriate selection of initial bistatic configuration can restrain from the appearance of the dead zone in common coverage. And the image results are obtained by frequency domain RD based on Method of Series Reversion (MSR). It is shown that GEO-UAV BiSAR has the high resolution ability.
Stability Problems for Chua System with One Linear Control
Authors: Arieşanu Camelia Pop
Published: April 2013

A Hamilton-Poisson realization and some stability problems for a dynamical system arisen from Chua system are presented. The stability and dynamics of a linearized smooth version of the Chua system are analyzed using the Hamilton-Poisson formalism. This geometrical approach allows to deduce the nonlinear stabilization near different equilibria.
Hypertrophic Cardiomyopathy Complicated by Pulmonary Edema in the Postpartum Period
Authors: Hanneman Kate , Nguyen Elsie T. , Crean Andrew M.
Published: April 2013

We report the case of a 42-year-old patient with hypertrophic cardiomyopathy (HCM) who presented to the emergency department with severe shortness of breath one week following uneventful cesarean delivery. Thoracic CT ruled out pulmonary embolus and confirmed pulmonary edema. Asymmetric interventricular septal thickening was clearly identified, demonstrating that the heart may be evaluated even on a non-ECG gated study. Acute pulmonary edema in the postpartum period is an unusual clinical presentation of HCM.
Extension of Axiomatic Design Principles for Multicriteria Decision Making Problems in Intuitionistic Fuzzy Environment
Authors: Li Ming
Published: April 2013

Axiomatic Design (AD) principles have been used to resolve the multicriteria decision making (MCDM) problems in engineering. With respect to MCDM problems in intuitionistic fuzzy environment, in which the criteria values take the form of intuitionistic fuzzy numbers, a new MCDM method is developed. Firstly, the approach proposed by Chen is extended to aggregate the decision makers’ opinions in intuitionistic fuzzy environment. Secondly, membership common area and nonmembership common area are derived from the membership probability density function and the nonmembership probability density function, respectively. Then the membership information content and nonmembership information content are obtained based on the basic ideal of axiomatic design principles. Afterwards, the score functionSand accuracy functionHin intuitionistic fuzzy sets are extended with the information content to compare the alternatives. The alternatives that have the lowest values of functions ofSandHare the best. Finally, a numerical example is used to illustrate the availability of the proposed method.
Optimization of Biodiesel Injection Parameters Based on Support Vector Machine
Authors: Shi Fuxi , Chen Jun , Xu Yang , Karimi Hamid Reza
Published: April 2013

For the running diesel engine, spray-atomization, mixed-combustion, and thermal-power conversion processes are inseparable, which causes difficulty to investigate atomization effect separately. This study was conducted to improve the atomization efficiency of the soybean fatty acid methyl ester (SFAME) in engine, to achieve the minimum effective specific fuel consumption in specific engine working conditions, the different injection parameters combination were explored on the influence of effective specific fuel consumption at elevated fule temperature. The effective specific fuel consumption prediction model was established based on support vector machine (SVM). With small samples, the intrinsic functional relationship was determined and the best injection parameters were validated under seven different experimental conditions. The study results have shown that the engine’s spray-thermal-power conversion process could be simulated accurately by using SVM. It will be more favorable to improve application effect of biodiesel in the engine to select the fuel temperature as injection parameters which influence atomization effect. Furthermore, using enumeration-verification methods to simulate the parameters might save a lot of resources as compared to the similar experiments.
Infarcted Adenomatoid Tumour of Epididymis: A Rare Case Report
Authors: Gupta A. , Livingston M. , Singh R. , Tansey D. , Solomon L.
Published: April 2013

Paratesticular tumours are pathologically rare. The vast majority are benign in nature with adenomatoid tumours representing the most common pathological entity. We present the case of a 32-year-old man, from the Indian subcontinent, who presented with a painful scrotal swelling sustained after trauma. The history suggested that the scrotal mass had been present for approximately 12 months, and a preliminary diagnosis of a haemorrhagic cyst caused by trauma was made. Initial management included scrotal support, analgesia, and a follow-up magnetic resonance imaging (MRI) scan. Subsequent imaging and then further histological analysis confirmed a partly necrotic/infarcted adenomatoid tumour of the right epididymis. After scrotal exploration and epididymectomy, the patient made a complete recovery, and, with the histological diagnosis, he was discharged with no further followup. The case is presented as a learning point in the identification and management of such pathologies.
Sequential Inhibitor Therapy in CML: In Vitro Simulation Elucidates the Pattern of Resistance Mutations after Second- and Third-Line Treatment
Authors: Bauer Robert C. , Sänger Jana , Peschel Christian , Duyster Justus , von Bubnoff Nikolas
Published: April 2013

Purpose: Dasatinib and nilotinib are active in imatinib-resistant chronic myelocytic leukemia (CML) and many patients undergo sequential treatment. We aimed at modeling sequential tyrosine kinase inhibitor (TKI) resistance in vitro to compare the sequences imatinib-nilotinib-dasatinib and imatinib-dasatinib-nilotinib.

Experimental Design: We designed an in vitro model for sequential TKI resistance in CML. Replicates of imatinib-resistant cell lines were treated with dasatinib or nilotinib. Second-line resistant replicates were exposed to third-line treatment.

Results: Growth of all replicates in all three lines of treatment was associated with T315I. However, T315I occurred with low abundance and did not increase during sequential treatment. Nilotinib second-line more often gave rise to sequential resistance compared with dasatinib due to pre-existing P-loop mutations, especially at suboptimal drug concentration. In contrast, mutations predisposing to dasatinib resistance such as F317C/V and V299L did not occur before dasatinib exposure. Nilotinib third-line did not overcome imatinib-dasatinib resistance due to pre-existing T315I or P-loop/V299L or P-loop/F317 exchanges. Dasatinib third-line suppressed imatinib-nilotinib–resistant replicates with residual sensitivity.

Conclusions: Sequential acquisition of BCR-ABL drug resistance mutations in CML might be underestimated. Resistance to sequential TKI monotherapy in vitro more often was associated with stepwise acquisition of drug-specific compound mutations compared with T315I. Pre-existing mutations strongly limited the activity of both third-line treatments, and the activity of nilotinib second-line in vitro critically depended on drug concentration. Clin Cancer Res; 19(11); 2962–72. ©2013 AACR.
Abstract A1: Long stress-induced non-coding transcripts (LSINCTs) and Cancer
Authors: Smith David I. , Jessica Silva M.
Published: April 2013

We previously utilized a whole genome tiling array to identify novel transcripts which had altered expression in response to the genotoxic effects of the tobacco carcinogen NNK. Our goal was to identify novel long stress-induced non-coding transcripts that could play important regulatory roles within cells and be important targets of alteration during cancer formation. Utilizing very stringent criteria, we examined all transcriptionally active regions that had altered expression in response to NNK treatment and identified a group of 12 transcripts, termed long stress-induced noncoding trancripts (LSINCTs). These novel long non-coding transcripts are between 2 and 4 Kb in length and do not contain any open reading frames or have homology to any mRNAs. 11 LSINCTs are intergenically located and one LSINCT is intragenically located within the genome. We found LSINCTs to be more highly expressed in more proliferative normal human tissues and to be frequently over-expressed in a number of different cancers including breast cancer. We studied one of these transcripts in greater detail, LSINCT5. We found LSINCT5 to be a polyadenylated nuclear transcript of exactly 2,647 base pairs. It is highly over-expressed in both breast and ovarian cancers and knocking down its' expression in either a breast or ovarian cancer cell line resulted in decreased cellular proliferation. We further compared gene expression in the breast cancer cell line, MCF7, before and after knocking down LSINCT5 expression and identified a number of genes whose expression changed more than 2-fold in response. Two of the genes identified were the gene for the major paraspeckle protein PSPC1, and another long non-coding transcript NEAT1. Paraspeckles are irregularly shaped compartments found in the nucleus' interchromatin space. Paraspeckles are RNA-protein structures formed by the interaction of NEAT1 and members of the Drosophila Behavior Human Splicing family of proteins including PSPC1. Paraspeckles appear to be critical to the control of gene expression through the nuclear retention of certain species of RNA. It is though that paraspeckles and their components play a role in controlling gene expression during a number of different cellular processes including differentiation, viral infection and stress responses. Using RNA fluorescent in situ hybridization, we found LSINCT5 to co-localize with NEAT1 within the nucleus. Hence, this novel stress-induced transcript may be another important long non-coding transcript that is involved in the normal cellular function of the paraspeckles.

Citation Format: David I. Smith, Silva M. Jessica. Long stress-induced non-coding transcripts (LSINCTs) and Cancer [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A1.
Abstract A17: MiR-15, miR-16 and miR-21 are direct transcriptional targets of E2F1 that limit E2F-induced proliferation and apoptosis
Authors: Ofir Matan , Ben-Shachar Berta , Hacohen Dalia , Ginsberg Doron
Published: April 2013

microRNAs (miRs) are small non-coding RNA molecules that have recently emerged as critical regulators of gene expression and are often deregulated in cancer. In particular, miRs encoded by the miR-15a, miR-16-1 cluster are frequently deleted in certain cancers and appear to act as tumor suppressors. Furthermore, miR-21 is over-expressed in a wide variety of human tumors and functions as an oncogene. Here, we provide evidence that miR-21, the miR-15a, miR-16-1 cluster and related miR-15b, miR-16-2 cluster comprise miRs regulated by E2F1, a pivotal transcription factor that can induce both proliferation and cell death. E2F1 is a critical downstream target of the tumor suppressor RB. The retinoblastoma pathway is often inactivated in human tumors resulting in deregulated E2F activity. We show that expression levels of the five mature miRs, miR-15a, miR-16-1, miR-15b, miR-16-2 and miR-21 are elevated upon activation of ectopic E2F1. Moreover, activation of endogenous E2Fs up-regulates expression of these miRs and endogenous E2F1 binds their respective promoters.

Importantly, we corroborate that miR-15a/b inhibits expression of cyclin E, a key direct transcriptional target of E2F pivotal for the G1/S transition. Similarly, miR-21 inhibits the expression of the pro-apoptotic tumor suppressor gene maspin, another direct transcriptional target of E2F. Therefore, our data suggest the existence of feed forward loops that modulate E2F activity and consist of E2F1; E2F1-regulated miRs -miR-15 and miR-21; and E2F1-regulated genes-cyclin E and maspin. In support of this, ectopic expression of miR-15 inhibits G1/S transition and, conversely, inhibition of miR-15 expression enhances E2F1-induced G1/S transition and up-regulation of cyclin E levels. Also, inhibition of miR-21 expression enhances E2F1-induced up regulation of maspin, as well as E2F1-mediated apoptosis.

In summary, our data identify miR-15, miR-16 and miR-21 as novel transcriptional targets of E2F that, in turn, modulate E2F activity.

Citation Format: Matan Ofir, Berta Ben-Shachar, Dalia Hacohen, Doron Ginsberg. MiR-15, miR-16 and miR-21 are direct transcriptional targets of E2F1 that limit E2F-induced proliferation and apoptosis [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A17.
Abstract A19: Profiling long noncoding RNA expression in cancer cells by real-time PCR arrays
Authors: Wu Fangting , Mo Yin-Yuan
Published: April 2013

Long noncoding RNAs (lncRNAs), as a new class of transcripts, have been recently shown to be pervasively transcribed in the genomes of human and mouse. In particular, accumulating evidence link the expression of lncRNAs to diverse human diseases, highlighting their potential as biomarkers and therapeutic targets. However, there are still many missing dots in our current understanding of lncRNA function. Thus, investigation of the expression patterns of lncRNAs is a crucial step to understanding of their roles in many model systems. To this end, we developed a qPCR based disease-related human lncRNA profiler, which allows for the quantification of differential expression of 83 individual lncRNAs among various experimental RNA samples. All 83 lncRNAs chosen for the array are based on publications and they are implicated in diseases ranging from neurodegeneration to cancer. The array plate also includes the house keeping genes and small RNA transcripts for normalization purposes. To determine the utility of this lncRNA profiler, we profiled their expression in breast cancer specimens compared to matched normal tissue and identified several lncRNAs are dysregulated, suggesting that lncRNAs may function as oncogenes or tumor suppressors. Gene expression profiling with this lncRNA profiler in a series of breast cell lines such as MCF10A, MCF7 and MDA-MB-231 cells identified differential expression patterns unique to each cell line, which may provide a basis for future analysis of these differentially expressed lncRNAs. Furthermore, we determined the effect of p53 on lncRNA expression in HCT-116 and MCF-7 cells. Initial testing indicated that at least three lncRNAs are induced by p53 in both cell lines. Experiments are underway to further characterize these p53 inducible lncRNAs. Together, these results suggest that disease-related human lncRNA profiler is a valuable research tool that can be tailored for specific needs in different research laboratories. The features of this profiler are easy, convenient, sensitive and specific. Accordingly it serves the first step toward the understanding of the role of lncRNAs in human diseases.

Citation Format: Fangting Wu, Yin-Yuan Mo. Profiling long noncoding RNA expression in cancer cells by real-time PCR arrays [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A19.
Abstract A21: MALAT-1 is essential for lung cancer metastasis in a novel human knockout model
Authors: Gutschner Tony , Eißmann Moritz , Hämmerle Monika , Stentrup Marion , Hildenbrandt Catherina , Groß Matthias , Zörnig Martin , Diederichs Sven
Published: April 2013

The highly conserved long non-coding RNA MALAT-1 (Metastasis-Associated in Lung Adenocarcinoma Transcript 1) had been discovered as a prognostic marker associated with poor survival and development of distant metastasis in lung adenocarcinoma. Since then, it has been found to be deregulated in numerous tumor entities and has been linked to splicing. However, its functional relevance in tumor cells remains to be elucidated. Knockdown models for MALAT1 have been described but suffer from insufficient silencing efficiency of the highly abundant, nuclear non-coding RNA (ncRNA).

In this project, we have developed a novel strategy to create ncRNA knockouts in human cancer cell lines. We have successfully used a synthetic Zinc Finger Nuclease engineered to target the 5′-region of MALAT1 to stably and biallelically integrate RNA-destabilizing elements into the genome of human lung cancer cells (A549). This approach resulted in a specific and more than 1000-fold silencing of MALAT1 in individual clones compared to a less than 5-fold silencing using siRNAs. Thus, this approach can be used to create functional knockouts of coding as well as non-coding genes also in human tumor cell lines allowing loss-of-function studies also of non-conserved ncRNAs in the future.

Phenotypically, the MALAT1-Knockout cells (KO) greatly differ from their parental cell line and wildtype clones (WT): Next to morphological changes, the migration of the KO cells is largely impaired as shown in scratch assays. In xenograft assays after i.v. injection, the KO cells form significantly fewer and smaller lung metastases than their WT counterparts. Since no large difference was observed after subcutaneous injection of the WT and the KO cells, this indicates a specific, active and essential function of MALAT1 in metastasis.

Exon microarrays of the WT and KO cell lines have revealed multiple migration- or metastasis-associated transcripts deregulated by loss of MALAT1 and hence potential target genes. These analyses also uncovered, that splicing is likely not the only functional mechanism of MALAT1 in the nucleus.

Taken together, we have developed a novel, highly effective approach for the knockout of genes that can be used for non-coding as well as coding RNAs in human tumor cells as well as cells from other species. Knockout of MALAT1 in human lung cancer cells revealed its essential function in metastasis as well as genes directly and indirectly targeted by MALAT1.

Citation Format: Tony Gutschner, Moritz Eißmann, Monika Hämmerle, Marion Stentrup, Catherina Hildenbrandt, Matthias Groß, Martin Zörnig, Sven Diederichs. MALAT-1 is essential for lung cancer metastasis in a novel human knockout model [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A21.
Abstract A24: Interaction of miR-106a~363Xpcl1 and p27Kip1 in thymopoiesis and lymphomagenesis
Authors: Kuppers Daniel , Schmitt Thomas , Clurman Bruce , Hwang Harry , Fero Matthew L.
Published: April 2013

The Xpcl1 (Kis2) gene encodes the miR-106a~363 miRNA cluster in mice. Although it was first identified as common integration site of the M-MuLV retrovirus in tumors from p27Kip1 (Cdkn1b) knockout mice, the nature of its interaction with cell cycle genes is uncertain. The miR-106a~363 miRNAs are highly conserved across species, implying an important role in normal biological processes. However, in the absence of a reported phenotype in knockout mice, its normal function is unclear. We examined miR-106a~363 expression in developing T lymphocytes and found that it is highly expressed in immature T-cells but is repressed at the CD4+/CD8+ double positive (DP) stage of thymopoiesis. Using a T cell specific Xpcl1 transgene, we found that forced expression of miR-106a~363 altered the distribution of T cell subsets. Xpcl1 repressed T-cell receptor expression and directly targeted the CD69 cell surface protein. This causes accumulation of DP thymocytes through complementary mechanisms.

We show that, in developing T-cells, p27Kip1 is differentially regulated due to variation in expression of FoxO family transcription factors. Likewise, p27 is upregulated at the transcriptional level by Xpcl1, which is surprising since this should have a tumor suppressor effect. Nonetheless, miR-106a~363 transgenic (Lx) mice developed spontaneous high grade T cell lymphomas. The combination of p27Kip1 deletion with miR-106a~363 overexpression, markedly accelerated lymphoma development, confirming cooperation of these two mutations. We conclude that deletion of p27Kip1 directly overcomes a tumor-suppressor activity of miR-106a~363, whereas the principal oncogenic function of the miRNA cluster remains unknown.

Citation Format: Daniel Kuppers, Thomas Schmitt, Bruce Clurman, Harry Hwang, Matthew L. Fero. Interaction of miR-106a~363Xpcl1 and p27Kip1 in thymopoiesis and lymphomagenesis [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A24.
Abstract A5: Cytotoxic Extracts of Ethnomedicinal Plants: Podophyllum peltatum (Mayapple) and Echinacea angustifolia
Authors: James Olena T. , Myles E. Lewis
Published: April 2013

Phytochemical and other natural compounds are historically established as cures for many ailments in rural America and other developing countries. Our lab is testing plants that show potential for inhibiting the growth of cancer cells. The family Asteraceae and the family Berberidacae are very well known for their values as medicinal plants. More specifically, the genus species Echinacea angustifolia and genus species Podophyllum peltatum commonly known as the Mayapple, both are noted for their unique medicinal purposes. The crude methanol extracts of these plants were tested against human metastatic cell lines representing breast (MCF-7), (BT-549) and colon (SW620) tissues. Trypan blue exclusion is used to determine viable cell counts. Alamar BlueTM and florescent analysis were used to evaluate the cytoxicity of the extracts. Analysis of the crude extracts of Echinacea angustifolia has shown on MCF-7, BT-549, and SW620 that most of them exhibit anticancer activity against the cell lines at the maximum concentration of 200 μg/mL. In particular, less diluted concentrations have demonstrated exceptional inhibitory effects with IC50 values ranging between 0.10–15 μg/mL, respectively. On the other hand, analysis of the crude extracts of Podophyllum peltatum (Mayapple) has shown on MCF-7, BT-549 and SW620 some anticancer activity against cell lines at the maximum concentration of 200 μg/mL. More specifically, the more diluted concentrations have demonstrated exceptional growth inhibition with IC50 values ranging between 2–8 μg/mL

Citation Format: Olena T. James, E. Lewis Myles. Cytotoxic Extracts of Ethnomedicinal Plants: Podophyllum peltatum (Mayapple) and Echinacea angustifolia [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A5.
Abstract A7: Roles of miRNAs in the switch of TGF-β from tumor suppressor to pro-oncogenic factor in cancer progression
Authors: Nagano Yoshiko , Lee Maxwell P. , Wakefield Lalage M.
Published: April 2013

TGF-β is a widely expressed pleiotropic growth factor that plays complex roles in tumorigenesis. TGF-β switches from tumor suppressor to pro-oncogenic factor during breast cancer progression. Our goal is to identify molecular determinants of the TGF-β switch. Since miRNA expression is deregulated in cancer, and TGF-β is reported to regulate expression and biogenesis of miRNAs, we hypothesize that miRNAs may contribute to this switch. To model the TGF-β switch, we used a series of human breast cancer cell lines derived from the spontaneously immortalized MCF10A human breast epithelial cell line. The model consists of 4 cell lines. In the MCF10A (M-I; normal), MCF10AT (M-II; premalignant) and MCF10CA1hcl4 (M-IIIcl4; low-grade tumor) lines, TGF-β is a tumor suppressor. In contrast, in the MCF10CA1a cell line (M-IV), which gives rise to aggressive metastatic tumors, the tumor suppressor activity is lost and TGF-β promotes metastasis. To test the possible involvement of miRNAs in the TGF-β switch, we compared expression profiles of miRNAs by Next Gen miRNA sequencing 1) basally in cultures of all four cell lines and 2) in M-IIIcl4 and M-IV with and without TGF-β treatment. A variety of patterns of change in basal miRNA expression with malignant progression were seen. We focused on miRNAs whose expression level differs basally between M-IIIcl4 and M-IV, since the switch occurs between these two cell lines. 46 miRNAs were expressed > 1.5 fold higher in M-IIIcl4 than in M-IV, whereas 29 miRNAs were expressed > 1.5 fold higher in M-IV than in M-IIIcl4. We selected several miRNAs whose expression was most different between the two cell lines, and examined whether these miRNAs regulate cellular responses to TGF-β. As an example, let-7i and let-7g were expressed higher in M-IV than in M-IIIcl4. Forced expression of these miRNAs partially reversed TGF-β-induced growth inhibition in M-IIIcl4. A similar strategy was applied to miRNAs whose regulation by TGF-β is different between M-IIIcl4 and M-IV. The results of these experiments will be discussed.

Citation Format: Yoshiko Nagano, Maxwell P. Lee, Lalage M. Wakefield. Roles of miRNAs in the switch of TGF-β from tumor suppressor to pro-oncogenic factor in cancer progression [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A7.
Abstract A8: Defining specific microRNAs that cooperatively regulate GRP78, an ER chaperone and signaling regulator, in cancer
Authors: Su Sheng-Fang , Lee Amy S. , Liang Gangning
Published: April 2013

GRP78, a major endoplasmic reticulum (ER) chaperone and signaling regulator, is commonly over-expressed in cancer and contributes to cancer cell proliferation, survival and tumorigenesis. Moreover, induction of GRP78 by a variety of anti-cancer drugs, including histone deacetylase inhibitors, confers chemo-resistance to cancer. Thus, targeting GRP78 to inhibit tumor survival and proliferation, sensitize tumor cells to chemotherapeutic drugs and consequently induce apoptosis serves as a promising avenue in anti-cancer therapy. As endogenous regulators, microRNAs (miRNAs) play important roles in modulating gene expression; therefore, we sought to identify miRNA(s) that target GRP78. We report that multiple miRNAs (miR-30d, miR-181a, miR-199-5p) commonly down-regulated in human cancer cell lines can repress GRP78 and lead to increased apoptosis in C42B prostate cancer cells. Luciferase reporter assays indicate that these miRNAs act as negative regulators of GRP78 by directly targeting the 3′ untranslated region of GRP78. Importantly, the combination of multiple miRNAs cooperated resulting in greater inhibition on GRP78 than individual miRNAs, suggesting the efficacy of tumor suppression by combined miRNAs. In addition, we showed that transfection of the multiple miRNAs in C42B cells increased sensitivity of cancer cells to trichostatin A (TSA), a histone deacetylase inhibitor, induced apoptosis, and inhibited cell proliferation and colony formation. Therefore, we suggest that delivery of combinations of specific miRNAs could be a novel anti-cancer approach capable of suppressing GRP78 and in turn inhibiting GRP78-mediated tumorigenesis.

Citation Format: Sheng-Fang Su, Amy S. Lee, Gangning Liang. Defining specific microRNAs that cooperatively regulate GRP78, an ER chaperone and signaling regulator, in cancer [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A8.
Abstract A9: De novo sequencing of circulating microRNAs identifies novel markers predicting clinical outcome of locally advanced breast cancer
Authors: Wu Xiwei , Somlo George , Palomares Melanie , Yen Yun , Rossi John , Gao Harry , Wang Shizhen Emily
Published: April 2013

Purpose: MicroRNAs (miRNAs) have been recently detected in the circulation of cancer patients, where they are associated with clinical parameters. Discovery profiling of circulating small RNAs has not been reported in breast cancer (BC), and was carried out in this study to identify blood-based small RNA markers of BC clinical outcome.

Patients and Methods: The pre-treatment sera of 42 stage II–III locally advanced and inflammatory BC patients who received neoadjuvant chemotherapy (NCT) followed by surgical tumor resection were analyzed for marker identification by deep sequencing all circulating small RNAs. An independent validation cohort of 26 stage II–III BC patients was used to assess the power of identified miRNA markers.

Results: More than 800 miRNA species were detected in the circulation, and exhibited patterns associated with the histopathological profiles of BC. Groups of circulating miRNAs differentially associated with ER/PR/HER2 status and inflammatory BC were identified. The relative levels of selected miRNAs measured by PCR showed consistency with their abundances determined by deep sequencing. Two circulating miRNAs, miR-375 and miR-122, exhibited strong correlations with clinical outcomes, including NCT response and relapse with metastatic disease. In the validation cohort, higher levels of circulating miR-122 specifically predicted metastatic recurrence in stage II–III BC patients.

Conclusion: Our study indicates that certain miRNAs can serve as potential blood-based biomarkers for NCT response, and that miR-122 prevalence in the circulation predicts BC metastasis in early-stage patients. These results may allow optimized chemotherapy treatments and preventive anti-metastasis interventions in future clinical applications.

Citation Format: Xiwei Wu, George Somlo, Melanie Palomares, Yun Yen, John Rossi, Harry Gao, Shizhen Emily Wang. De novo sequencing of circulating microRNAs identifies novel markers predicting clinical outcome of locally advanced breast cancer [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A9.