bluuebunny/crossref_metadata_2025 huggingface dataset:
Systems Pharmacology-Based Approach to Comparatively Study the Independent and Synergistic Mechanisms of Danhong Injection and Naoxintong Capsule in Ischemic Stroke Treatment
Authors: Zhu Junfeng , Yi Xiaojiao , Zhang Yiwen , Pan Zongfu , Zhong Like , Huang Ping
Published: February 2019

To provide evidence for the better clinical use of traditional Chinese medicine preparations (TCMPs), comparison of the pharmacological mechanisms between TCMPs with similar therapeutic effect is necessary. However, methodology for dealing with this issue is still scarce. Danhong injection (DHI) and Naoxintong capsule (NXT) are representative TCMPs for ischemic stroke (IS) treatment, which are also frequently used in combination. Here they were employed as research objects to demonstrate the feasibility of systems pharmacology approach in elucidation of the independent and combined effect of TCMPs. By incorporating chemical screening, target prediction, and network construction, a feasible systems pharmacology model has been established to systematically uncover the underlying action mechanisms of DHI, NXT, or their pair in IS treatment. Systematic analysis of the created TCMP-Compound-Target-Disease network revealed that DHI and NXT shared common targets such as PTGS2, F2, ADRB1, IL6, ALDH2, and CCL2, which were involved in the vasomotor system regulation, blood-brain barrier disruption, redox imbalance, neurotrophin activity, and brain inflammation. In comparative mechanism study, the merged DHI/NXT-IS PPI network and pathway enrichment analysis indicated that DHI and NXT exerted the therapeutic effects mainly through immune system and VEGF signaling pathways. Meanwhile, they had their own unique pathways, e.g., calcium signaling pathway for DHI and gap junction for NXT. While for their synergistic mechanism, DHI and NXT participated in chemokine signaling pathway, T cell receptor signaling pathway, VEGF signaling pathway, gap junction, and so on. Our study provided an optimized strategy for dissecting the different and combined effect of TCMPs with similar actions.
Citrus Alkaline Extract Delayed the Progression of Pulmonary Fibrosis by Inhibiting p38/NF-κB Signaling Pathway-Induced Cell Apoptosis
Authors: Wu Qi , Zhou Yao , Zhou Xian-mei
Published: February 2019

Objective. To investigate the intervention effect and functioning mechanism of citrus alkaline extract (CAE) on bleomycin- (BLM-) induced pulmonary fibrosis in mice. Methods. 42 C57BL/6 male mice were assigned randomly to the normal group, model group, low (16mg/kg), medial (32mg/kg) and high (64mg/kg) CAE dosage groups, prednisone group (6mg/kg), and pirfenidone group (100mg/kg), respectively. One day after model construction, intragastric administration was applied to the mice once a day for 28 days and then killed. Body weights of mice were recorded. Their pulmonary tissues were subjected to HE staining and Masson’s staining and then their degree of pulmonary alveolitis as well as pulmonary fibrosis was scored. Contents of hydroxyproline (HYP) and prostaglandin E2 (PGE2) in pulmonary tissues and levels of interleukin-17 (IL-17) in serum and bronchoalveolar lavage fluid (BALF) were determined by ELISA method. Expression of collagen I, collagen III, and Prosurfactant protein C (Pro-SPC) proteins in pulmonary tissue were measured immunohistochemically and that of nuclear transcription factor κB (NF-κB) and vimentin was determined by the immunofluorescence method. Apoptosis of pulmonary tissue was tested by the Tunel staining method, while the expression of MAPK-related protein was recorded by Western Blot assay. Results. After CAE treatment, the body weight, PGE2 level, and Pro-SPC protein expression of pulmonary fibrosis mice were increased, while the score of pulmonary alveolitis and pulmonary fibrosis, levels of HYP and cell apoptosis, IL-17 contents of serum and BALF in pulmonary tissues, and expression of collagen I, collagen III, vimentin, NF-κB, and p-p38 were reduced. Conclusion. CAE effectively delayed the progression of BLM-induced pulmonary fibrosis in pulmonary fibrosis mice and a possible mechanism is the inhibition of cell apoptosis of NF-κB/p38-mediated signaling pathway.
Chemokine Receptor 2 (CXCR2) Gene Variants and Their Association with Periodontal Bacteria in Patients with Chronic Periodontitis
Authors: Kavrikova Denisa , Borilova Linhartova Petra , Lucanova Svetlana , Poskerova Hana , Fassmann Antonin , Izakovicova Holla Lydie
Published: February 2019

Periodontitis, an inflammatory disease caused by subgingival Gram-negative (G-) bacteria, is linked with loss of the connective tissue and destruction of the alveolar bone. In the regulation of inflammatory response, chemokine receptor 2 (CXCR2), a specific receptor for interleukin-8 and neutrophil chemoattractant, plays an important role. The first aim of this study was to investigate theCXCR2gene variability in chronic periodontitis (CP) patients and healthy nonperiodontitis controls in the Czech population. The second aim was to find a relation betweenCXCR2gene variants and the presence of periodontal bacteria. A total of 500 unrelated subjects participated in this case-control study. 329 CP patients and 171 healthy nonperiodontitis controls were analyzed using polymerase chain reaction techniques for three single-nucleotide polymorphisms (SNPs): +785C/T (rs2230054), +1208T/C (rs1126579), and +1440A/G (rs1126580). A DNA microarray detection kit was used for the investigation of the subgingival bacterial colonization, in a subgroup of CP subjects (N=162). No significant differences in allele, genotype, haplotype, or haplogenotype frequencies ofCXCR2gene variants between patients with CP and healthy controls (P>0.05) were determined. Nevertheless,Aggregatibacter actinomycetemcomitanswas detected more frequently in men positive for the C allele of theCXCR2+785C/T polymorphism (61.8% vs. 41.1%,P<0.05;OR=2.31, 95%CI=1.03-5.20) and for the T allele of theCXCR2+1208C/T variant (61.8% vs. 38.9%,P<0.05;OR=2.54, 95%CI=1.13-5.71). In contrast, no statistically significant associations ofCXCR2variants with seven selected periodontal bacteria were found in women. Although none of the investigated SNPs in theCXCR2gene was associated with CP, theCXCR2gene variants can be associated with subgingival colonization of G- bacteria in men with CP in the Czech population.
A Feature Extraction Method of Hybrid Gram for Malicious Behavior Based on Machine Learning
Authors: Zhao Yuntao , Bo Bo , Feng Yongxin , Xu ChunYu , Yu Bo
Published: February 2019

With explosive growth of malware, Internet users face enormous threats from Cyberspace, known as “fifth dimensional space.” Meanwhile, the continuous sophisticated metamorphism of malware such as polymorphism and obfuscation makes it more difficult to detect malicious behavior. In the paper, based on the dynamic feature analysis of malware, a novel feature extraction method of hybrid gram (H-gram) with cross entropy of continuous overlapping subsequences is proposed, which implements semantic segmentation of a sequence of API calls or instructions. The experimental results show the H-gram method can distinguish malicious behaviors and is more effective than the fixed-length n-gram in all four performance indexes of the classification algorithms such as ID3, Random Forest, AdboostM1, and Bagging.
TNF-α Downregulation Modifies Insulin Receptor Substrate 1 (IRS-1) in Metabolic Signaling of Diabetic Insulin-Resistant Hepatocytes
Authors: Alipourfard Iraj , Datukishvili Nelly , Mikeladze Davit
Published: February 2019

One of the major mechanisms of hyperglycemia in type 2 diabetes is insulin resistance (IR) which can induce free fatty acids like palmitate. In hepatic cell, as an insulin target tissue, insulin resistance can be stimulated by inflammatory cytokine TNF-α. The interaction of intracellular TNF-α signal with the insulin signaling pathway is not well identified. Hence, we aimed to investigate the effect of TNF-α elimination on the diabetic model of palmitate-induced insulin-resistant hepatocytes (HepG2). The changes of phosphorylation rate in IRS-1 protein are determined to know the effect of TNF-α on this key protein of the insulin signaling pathway. HepG2 cells were treated with 0.5 Mm palmitate, and TNF-α gene knockdown was performed by shRNA-mediated technique. Western blot analysis was used to evaluate the phosphorylated activity of the insulin signaling pathway. Palmitate-induced IR could increase TNF-α protein expression 1.2-, 2.78-, and 2.25-fold compared to the control cells at times of 8 h, 16 h, and 24 h, respectively. TNF-α expression in downregulated cells transfected with shRNA-TNF-α is approximately 47.0% of normal cells and 49.0% in the case of scrambled cells. IRS-1 phosphorylation in TNF-α-downregulated and stimulated cells with 100 nM insulin, after treatment and in the absence of palmitate, was 45% and 29% higher than the normal cells. These data support the evidence that TNF-α downregulation strategy contributes to the improvement of IRS-1 phosphorylation after insulin stimulation and insulin response in HepG2 liver cells.
Efficient Prediction of Network Traffic for Real-Time Applications
Authors: Iqbal Muhammad Faisal , Zahid Muhammad , Habib Durdana , John Lizy Kurian
Published: February 2019

Accurate real-time traffic prediction is required in many networking applications like dynamic resource allocation and power management. This paper explores a number of predictors and searches for a predictor which has high accuracy and low computation complexity and power consumption. Many predictors from three different classes, including classic time series, artificial neural networks, and wavelet transform-based predictors, are compared. These predictors are evaluated using real network traces. Comparison of accuracy and cost, both in terms of computation complexity and power consumption, is presented. It is observed that a double exponential smoothing predictor provides a reasonable tradeoff between performance and cost overhead.
Synthesis and Characterization of Silver-Containing Nanocomposites Based on 1-Vinyl-1,2,4-triazole and Acrylonitrile Copolymer
Authors: Pozdnyakov Alexander S. , Emel’yanov Artem I. , Kuznetsova Nadezhda P. , Ermakova Tamara G. , Korzhova Svetlana A. , Khutsishvili Spartak S. , Vakul’skaya Tamara I. , Prozorova Galina F.
Published: February 2019

Insoluble silver-containing nanocomposites based on 1-vinyl-1,2,4-triazole and acrylonitrile copolymer was synthesized and characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, atomic absorption, X-ray diffraction, and thermogravimetric data analysis. The silver content of the nanocomposites varied from 8.0 to 23.5% (the silver content depends on the ratio of the copolymer-silver salt). Synthesis of silver nanocomposites was carried out by thermal treatment of a polymer complex of copolymer-silver salt without the use of an additional reducing agent. X-ray diffraction was used to control the formation of silver nanoparticles at different temperatures (210°С, 220°С, and 250°С). Synthesis of nanocomposites shows a developed porous structure with a channel diameter in most cases of 1-4 microns. The EPR monitoring of the nanoparticle germination and growth reveals that the generation of the nanocomposites proceeds via a stage of complex formation between the polymeric matrix and silver ions. The synthesized nanocomposites possess paramagnetic properties and show a narrow EPR symmetric signal with the concentration of unpaired electrons of 1020 spin/g, a g-factor 2.0050, and signal width 5-6 Gs. Thermal stability of silver-containing nanocomposites is up to 270-320°C.
ST-Elevation Myocardial Infarction (STEMI) in a Patient with Moyamoya Disease
Authors: Livesay James , Johnson Jeffrey
Published: February 2019

Moyamoya disease is a rare condition that is primarily reported in Asian populations, characterized by stenoocclusive intracranial angiopathy with small, fragile, and multiple collateral vessel formation. Extracranial complications, mainly abnormalities within the renal vasculature, have been described; however, there are very few case reports of cardiovascular complications in patients with Moyamoya disease. We report a 26-year-old Caucasian female with known Moyamoya disease who presented with both typical and atypical chest pain, mimicking symptoms of a previous non-ST-elevation myocardial infarction. Approximately six months prior to the current hospital admission, she underwent coronary angiography requiring percutaneous coronary intervention (PCI) with two drug-eluting stents to the right coronary artery (RCA) for a critical stenosis. Despite medical management, our patient developed inferior lead ST-elevations leading to a repeat left heart catheterization which showed clinically significant stenosis of the first obtuse marginal branch. Development of significant coronary artery stenosis in a short period of time demonstrates the clinical significance of minimal atherosclerosis in the setting of underlying fibrocellular thickening as seen in patients with Moyamoya disease. Clinicians need to be aware of the possibility of coronary involvement in addition to intracranial vascular complications in patients with Moyamoya disease and take appropriate measures to prevent or delay the development of atherosclerosis in these arteries.
Nanoencapsulation of Curcumin and Its Protective Effects against CCl4-Induced Hepatotoxicity in Mice
Authors: Li Jian , Niu Ruixu , Dong Lixin , Gao Liming , Zhang Jie , Zheng Yue , Shi Ming , Liu Zhiwei , Li Kun
Published: February 2019

Curcumin is a natural phenolic compound extracted from the herbCurcuma longaL. rhizome and has received much attention on account of its biological properties. However, its poor solubility and low bioavailability limit its use. The purpose of this study was to synthesize curcumin-loaded nanoliposomes (Cur-NLs) to improve their bioavailability and evaluate the hepatoprotective effect of Cur-NLs against tetrachloromethane- (CCl4-) induced acute liver injury in mice. We prepared Cur-NLs by thin film dispersion method, and the characterizations of Cur-NLs were measured by transmission microscope, laser particle size analyzer, infrared spectrometer, and X-ray diffraction. After 14 days pretreatment of Cur-NLs, free curcumin, silybin, or PBS, the models of acute liver injury were established by CCl4intraperitoneal injection in mice. The organ index, biochemical liver function parameters, histopathology, and antioxidant enzyme activities of liver tissues were measured further to evaluate the protective effects of Cur-NLs on liver injury. Compared with the CCl4model control group, pretreatment of Cur-NLs effectively reduced the serum levels of ALT, AST, and ALP and attenuated the hepatic necrosis induced by CCl4intoxication. Furthermore, Cur-NL pretreatment remarkably exhibited decreased MDA level and increased SOD, GPx, and CAT activities compared to CCl4model control group. Compared with the free curcumin group, the Cur-NLs also showed a better hepatoprotective effect. These observations imply that Cur-NLs act as a promising hepatoprotective agent in reducing liver oxidative stress produced by different stress factors.
Automatic Nonnutritive Suck Waveform Discrimination and Feature Extraction in Preterm Infants
Authors: Liao Chunxiao , Rosner Austin O. , Maron Jill L. , Song Dongli , Barlow Steven M.
Published: February 2019

Background and Objective: The emergence of the nonnutritive suck (NNS) pattern in preterm infants reflects the integrity of the brain and is used by clinicians in the neonatal intensive care unit (NICU) to assess feeding readiness and oromotor development. A critical need exists for an integrated software platform that provides NNS signal preprocessing, adaptive waveform discrimination, feature detection, and batch processing of big data sets across multiple NICU sites. Thus, the goal was to develop and describe a cross-platform graphical user interface (GUI) and terminal application known as NeoNNS for single and batch file time series and frequency-domain analyses of NNS compression pressure waveforms using analysis parameters derived from previous research on NNS dynamics.Methods. NeoNNS was implemented with Python and the Tkinter GUI package. The NNS signal-processing pipeline included a low-pass filter, asymmetric regression baseline correction, NNS peak detection, and NNS burst classification. Data visualizations and parametric analyses included time- and frequency-domain view, NNS spatiotemporal index view, and feature cluster analysis to model oral feeding readiness.Results. 568 suck assessment files sampled from 30 extremely preterm infants were processed in the batch mode (<50 minutes) to generate time- and frequency-domain analyses of infant NNS pressure waveform data. NNS cycle discrimination and NNS burst classification yield quantification of NNS waveform features as a function of postmenstrual age. Hierarchical cluster analysis (based on the Tsfresh python package and NeoNNS) revealed the capability to label NNS records for feeding readiness.Conclusions. NeoNNS provides a versatile software platform to rapidly quantify the dynamics of NNS development in time and frequency domains at cribside over repeated sessions for an individual baby or among large numbers of preterm infants at multiple hospital sites to support big data analytics. The hierarchical cluster feature analysis facilitates modeling of feeding readiness based on quantitative features of the NNS compression pressure waveform.
Thiazole Based Carbohydrazide Derivatives as α-Amylase Inhibitor and Their Molecular Docking Study
Authors: Taha Muhammad , Irshad Maryam , Imran Syahrul , Rahim Fazal , Selvaraj Manikandan , Almandil Noor Barak , Mosaddik Ashik , Chigurupati Sridevi , Nawaz Faisal , Ismail Nor Hadiani , Ibrahim Mohamed
Published: February 2019

In this study we are going to present thiazole based carbohydrazide in search of potent antidiabetic agent as α-amylase inhibitors. Thiazole based carbohydrazide derivatives 1-25 have been synthesized, characterized by 1HNMR, 13CNMR, and EI-MS, and evaluated for α-amylase inhibition. Except compound 11 all analogs showed α-amylase inhibitory activity with IC50 values from 1.709 ± 0.12 to 3.049 ± 0.25 μM against the standard acarbose (IC50 = 1.637 ± 0.153 μM). Compounds 1, 10, 14, and 20 exhibited outstanding inhibitory potential with IC50 value 1.763 ± 0.03, 1.747 ± 0.20, 1.709 ± 0.12, and 1.948 ± 0.23 μM, respectively, compared with the standard acarbose. Structure activity relationships have been established for the active compounds. To get an idea about the binding interaction of the compounds, molecular docking studies were done.
Diffusion and Friction Dynamics of Probe Molecules in Liquid n-Alkane Systems: A Molecular Dynamics Simulation Study
Authors: Lee Song Hi
Published: February 2019

We present a molecular dynamics simulation study of the probe diffusion and friction dynamics of Lennard-Jones particles in a series of liquid n-alkane systems from C12 up to C400 at 318 K, 418 K, 518 K, and 618 K, to investigate the power law dependence of self-diffusion of polymer liquids on their molecular weights. Two LJ particles MY1 with a mass of 114 g/mol and MY2 with a mass of 225 g/mol are used as probes to model methyl yellow. We observed that a clear transition in the power law dependence of n-alkane self-diffusion on the molecular weight (M) of n-alkane, Dself∼M−γ, occurs in the range C120∼C160 at temperatures of 318 K, 418 K, and 518 K, corresponding to a crossover from the “oligomer” to the “Rouse” regime. We also observed that a clear transition in the power law dependence of the diffusion coefficient DMY2 on the molecular weight (M) of n-alkane, DMY2∼M−γ, occurs at low temperatures. The exponent γ for DMY2 shows a sharp transition from 1.21 to 0.52 near C36 at 418 K and from 1.54 to 0.60 near C36 at 318 K. However, no such transition is found for the probe molecule MY2 at temperatures of 518 K and 618 K and for MY1 probe at temperatures of 418 K, 518 K, and 618 K, but the power law exponent γ for MY1 at 318 K shows instead a linear or a rather slow transition. The dependence of the probe diffusion (DMY2) on the matrix molecular weight (M) reflects a significant change of the matrix dynamics associated with the probe diffusion: a crossover from the “solvent-like” to the “oligomer” regime. As the molecular weight of n-alkane increases, the ratio of Dself/DMY2 becomes less than 1 and the probe molecules encounter, in turn, two different microscopic frictions depending on MMY/Mmatrix and the temperature. It is believed that a reduction in the microscopic friction on the probe molecules that diffuse at a rate faster than the solvent fluctuations leads to large deviations of slope from the linear dependence of the friction of MY2 on the chain length of the n-alkane at 318 K and 418 K.
Genetic Ancestry–dependent Differences in Breast Cancer–induced Field Defects in the Tumor-adjacent Normal Breast
Authors: Nakshatri Harikrishna , Kumar Brijesh , Burney Heather N. , Cox Mary L. , Jacobsen Max , Sandusky George E. , D'Souza-Schorey Crislyn , Storniolo Anna Maria V.
Published: February 2019

Purpose:

Genetic ancestry influences evolutionary pathways of cancers. However, whether ancestry influences cancer-induced field defects is unknown. The goal of this study was to utilize ancestry-mapped true normal breast tissues as controls to identify cancer-induced field defects in normal tissue adjacent to breast tumors (NATs) in women of African American (AA) and European (EA) ancestry.

Experimental Design:

A tissue microarray comprising breast tissues of ancestry-mapped 100 age-matched healthy women from the Komen Tissue Bank (KTB) at Indiana University (Indianapolis, IN) and tumor-NAT pairs from 100 women (300 samples total) was analyzed for the levels of ZEB1, an oncogenic transcription factor that is central to cell fate, mature luminal cell–enriched estrogen receptor alpha (ERα), GATA3, FOXA1, and for immune cell composition.

Results:

ZEB1+ cells, which were localized surrounding the ductal structures of the normal breast, were enriched in the KTB-normal of AA compared with KTB-normal of EA women. In contrast, in EA women, both NATs and tumors compared with KTB-normal contained higher levels of ZEB1+ cells. FOXA1 levels were lower in NATs compared with KTB-normal in AA but not in EA women. We also noted variations in the levels of GATA3, CD8+ T cells, PD1+ immune cells, and PDL1+ cell but not CD68+ macrophages in NATs of AA and EA women. ERα levels did not change in any of our analyses, pointing to the specificity of ancestry-dependent variations.

Conclusions:

Genetic ancestry–mapped tissues from healthy individuals are required for proper assessment and development of cancer-induced field defects as early cancer detection markers. This finding is significant in light of recent discoveries of influence of genetic ancestry on both normal biology and tumor evolution.
Abstract A17: Metronomic topotecan causes MYCN inactivation and impedes tumor growth selectively in MYCN-amplified neuroblastoma cells in vitro and in vivo by therapy-induced senescence
Authors: Taschner-Mandl Sabine , Gerber Teresa , Schwarz Magdalena , Blaha Johanna , Kauer Maximilian , Kromp Florian , Frank Nelli , Rifatbegovic Fikret , Weiss Tamara , Ladenstein Ruth , Hohenegger Martin , Ambros Inge M. , Ambros Peter F.
Published: February 2019

Background: Poor prognosis and frequent relapses are major challenges for patients with high-risk neuroblastoma (NB), especially when tumors show MYCN amplification. High-dose chemotherapy triggers apoptosis, necrosis, and senescence, a cellular stress response leading to permanent proliferative arrest and a typical senescence-associated secretome (SASP). SASP components reinforce growth arrest and act in an immune-stimulatory fashion, while others are tumor promoting. Studies on low-dose long-term, i.e., metronomic, treatment schedules have demonstrated effects on the tumor microenvironment, but have not comprehensively analyzed cellular responses in tumor cells.

Methods and Results: We evaluated efficacy and mode of action of low-dose metronomic treatment schedules in in vitro and in vivo NB models. Importantly, by using the secretome as a discriminator for beneficial versus adverse effects of senescence, drugs with a tumor-inhibiting SASP were identified. Our preclinical work has demonstrated that low-dose long-term topotecan (TPT) treatment (0.1 mg/kg/d, i.p., daily over 15 weeks) improves survival in mouse NB xenografts and leads to long-term cure in 40% of mice. TPT induces a tumor-inhibiting type of senescence, accompanied by changes in the tumor transcriptome and secretome, specifically in MYCN-amplified NB cells. As senescent NB cells showed a reduced MYCN copy number and strongly downregulated MYCN expression, we investigated the mechanism of MYCN inactivation. This showed that upon senescence MYCN is epigenetically silenced and gene copies are recruited to the nuclear periphery where changes in the nuclear lamina composition take place.

Conclusion: This new mode of action of metronomic TPT treatment, i.e., promoting a tumor-inhibiting type of senescence selectively in MYCN-amplified neuroblastoma, is clinically relevant as metronomic regimens are increasingly implemented in therapy protocols of various cancer entities and are considered as a feasible maintenance treatment option with moderate adverse event profiles.

Citation Format: Sabine Taschner-Mandl, Teresa Gerber, Magdalena Schwarz, Johanna Blaha, Maximilian Kauer, Florian Kromp, Nelli Frank, Fikret Rifatbegovic, Tamara Weiss, Ruth Ladenstein, Martin Hohenegger, Inge M. Ambros, Peter F. Ambros. Metronomic topotecan causes MYCN inactivation and impedes tumor growth selectively in MYCN-amplified neuroblastoma cells in vitro and in vivo by therapy-induced senescence [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr A17.
Abstract B32: A novel role for the EWS portion of EWS/FLI in binding GGAA-microsatellites required for oncogenic transformation in Ewing sarcoma
Authors: Johnson K. M. , Lessnick S. L.
Published: February 2019

Objective: Ewing sarcoma is a pediatric bone malignancy initiated by a t(11;22) chromosomal translocation that produces the EWS/FLI oncoprotein. EWS/FLI transcriptionally activates and represses its target genes to mediate oncogenic reprogramming. Expression of its upregulated targets correlates with EWS/FLI binding to associated GGAA-microsatellites, which show length polymorphisms. These microsatellite polymorphisms may critically affect EWS/FLI-responsiveness of key gene targets. For example, NR0B1 is necessary for EWS/FLI mediated oncogenic transformation, and we found a “sweet spot” of 18-26 repeat length as optimal for EWS/FLI mediated transcriptional activity at NR0B1 through clinical observations and in vitro studies. The mechanism underlying this optimal length is unknown.

Methods: We explored the stoichiometry and binding affinity of EWS/FLI for different GGAA-repeat lengths through biochemical studies, including fluorescence polarization, ChIP-seq, and RNA-seq, combined with bioinformatics analysis. Additionally, use of mutant constructs of EWS/FLI has been critical for elucidating particular binding behavior of EWS/FLI at different microsatellite repeat lengths.

Results: Our biochemical studies, using recombinant Δ22 (a version of EWS/FLI containing only the FLI portion) demonstrate a stoichiometry of one Δ22-monomer binding to every two consecutive GGAA-repeats on shorter microsatellite sequences. Surprisingly, the affinity for Δ22 binding to GGAA-microsatellites significantly decreased, and ultimately became unmeasurable, when the size of the microsatellite was increased to the “sweet spot” length. In contrast, a fully functional EWS/FLI mutant (Mut9, which retains approximately half of the EWS portion of the fusion) showed low affinity for smaller GGAA-microsatellites, but instead significantly increased its affinity at “sweet spot” microsatellite lengths. Single-gene ChIP and genome-wide ChIP-seq and RNA-seq studies extended these findings to the in vivo setting.

Conclusion: Together, these data reveal an unexpected novel role for the EWS portion of the EWS/FLI fusion in DNA binding. Additionally, our data suggest a length-dependent biochemical mechanism for EWS/FLI binding and transcriptional regulation at GGAA-microsatellites.

Citation Format: K. M. Johnson, S. L. Lessnick. A novel role for the EWS portion of EWS/FLI in binding GGAA-microsatellites required for oncogenic transformation in Ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr B32.
Abstract IA03: Subgroup-specific enhancer hijacking in medulloblastoma
Authors: Northcott Paul A.
Published: February 2019

In recent years, next-generation sequencing has uncovered the spectrum of driver genes and molecular processes contributing to most cancer entities, including pediatric cancer. This holds true for medulloblastoma (MB), the most common malignant childhood brain tumor. A multitude of genomic studies conducted on large patient cohorts have culminated in the recognition of at least four molecular and clinically distinct MB subgroups: WNT, SHH, Group 3, and Group 4. Moreover, comprehensive sequencing efforts have reported the prevalence and subgroup distribution of somatically altered genes in MB, informing tumor biology and suggesting rational candidates for molecularly targeted therapy. In contrast to WNT and SHH subgroup MB, which are typically characterized by mutations/copy-number alterations leading to aberrant signaling of the WNT and SHH pathways, respectively, Groups 3 and 4 harbor a paucity of recurrently mutated genes and exhibit remarkable heterogeneity in their molecular landscapes. Looking beyond somatic gene-level mutations and focusing our analyses on recurrent structural variants (SVs) in Group 3 and Group 4, we recently discovered a series of atypical SVs converging on chr9q34 that coincided with aberrant induction of GFI1B expression, a transcription factor normally active in the hematopoietic system. Meticulous mapping of SV breakpoints and subsequent integration with DNA methylation and histone ChIP-seq datasets implicated misappropriation of highly active enhancers secondary to the observed SV, a mechanism we termed “enhancer hijacking.” In light of this surprising discovery, we have now extended our analyses, including the development of new computational pipelines, to systematically explore the extent of enhancer hijacking in MB and other pediatric brain tumors. Emerging results from these efforts will be presented, including strategies being employed to validate aberrant enhancer-promoter interactions in MBs with presumed enhancer hijacking, as well as functional approaches we are using to engineer these alterations in relevant cellular systems.

Citation Format: Paul A. Northcott. Subgroup-specific enhancer hijacking in medulloblastoma [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr IA03.
Abstract IA08: Epigenetic mechanisms of resistance to targeted therapy
Authors: George Rani E.
Published: February 2019

Targeted cancer therapies, such as inhibitors of the ALK receptor tyrosine kinase, have yielded promising outcomes in patients whose tumors aberrantly express ALK, yet the development of resistance invariably limits their efficacy. To address this issue, we have investigated potential mechanisms of resistance to ALK inhibitors in neuroblastoma cells expressing ALK point mutations. In tumor cells expressing the ALKF1174L mutant kinase in the absence of MYCN amplification, we observed that resistance to ALK inhibitors develops through activation of MAPK signaling and induction of epithelial-to-mesenchymal transition. By contrast, in tumor cells expressing the same mutation but with concomitant MYCN overexpression due to genomic amplification, resistance arises through a different mechanism, characterized by extensive chromatin remodeling and conversion to a stem cell-like state. I will describe our latest insights into the molecular details underlying this latter mechanism with emphasis on transcriptional regulators.

Citation Format: Rani E. George. Epigenetic mechanisms of resistance to targeted therapy [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr IA08.
Abstract IA15: SWI/SNF (BAF) chromatin remodeling complexes are frequently mutated in cancer: Mechanisms and vulnerabilities
Authors: Roberts Charles W. M.
Published: February 2019

Data emerging over the last several years implicate SWI/SNF (BAF) chromatin remodeling complexes as a major tumor suppressor as frequent inactivating mutations in at least nine different SWI/SNF subunits, collectively identified in 20% of all cancers. These include recurrent mutations of ARID1A (BAF250a) in ovarian, endometrioid, bladder, stomach, colorectal, and pancreatic cancers and neuroblastoma; of the BRG1 (SMARCA4) subunit in medulloblastomas and non-small cell lung cancers; of the PBRM1 (BAF180) subunit in renal carcinomas; of the ARID2 subunit in hepatocellular, lung, and pancreas carcinomas as well as melanomas; and of the BRD7 subunit in breast cancers. The SWI/SNF complex includes both core and lineage-specific subunits and utilizes the energy of ATP to modulate chromatin structure and regulate transcription. My laboratory began studying the SWI/SNF complex when SNF5 (SMARCB1/INI1/BAF47) became the first SWI/SNF subunit linked to tumor suppression over fifteen years ago when it was found to be biallelically inactivated in nearly all cases of a highly aggressive type of pediatric cancer called malignant rhabdoid tumor (MRT). Despite the extremely aggressive and lethal nature of MRT, we have shown that these cancers are diploid and have remarkably simple genomes. We now study the complex using mouse models, cell lines, and primary human tumor samples. Insights into the normal function of SWI/SNF complexes, the mechanisms by which mutation of the complexes drive cancer formation, and potential therapeutic vulnerabilities created by mutation of the complex will be presented, including our recent efforts that identify a central role for SWI/SNF complexes in enhancer regulation.

Citation Format: Charles W. M. Roberts. SWI/SNF (BAF) chromatin remodeling complexes are frequently mutated in cancer: Mechanisms and vulnerabilities [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr IA15.
Abstract A023: Regulation of ERG function in prostate cells by phosphorylation and interaction with Ewing’s sarcoma breakpoint protein EWS
Authors: Kedage Vivekananda , Nicholas Taylor R. , Strittmatter Brady G. , Selvaraj Nagarathinam , Budka Justin A. , Jerde Travis J. , Hollenhorst Peter C.
Published: February 2019

More than one-half of prostate tumors have a chromosomal rearrangement that results in the overexpression of an oncogenic ETS family transcription factor. The most common fusion, TMPRSS2/ERG, results in expression of ERG, a protein that is not normally expressed in prostate epithelia. When ERG is expressed in prostate cells it is thought to bind to enhancer elements and regulate gene expression by recruiting transcriptional coactivators and/or corepressors. We have recently shown that the EWS protein acts as a coactivator for ERG in prostate cells and this interaction is required for ERG-mediated xenograft tumor growth. Interestingly, the EWS interaction may be the key requirement that separates oncogenic from nononcogenic ETS factors, as EWS only interacts with four ETS family members involved in prostate cancer gene rearrangements (ERG, ETV1, ETV4, and ETV5), but not with other ETS family members. This ETS/EWS interaction also indicates a common molecular mechanism involved in prostate cancer and Ewing’s sarcoma, a cancer caused by gene fusions that express chimeric EWS/ETS proteins. ERG also interacts with corepressors such as EZH2, a subunit of PRC2. We have recently found that the interaction between ERG and EZH2/PRC2 is regulated by a series of phosphorylation events on ERG. The MAP kinase ERK can bind a high-affinity docking sequence in ERG, resulting in phosphorylation of a nearby serine, S215. This phosphorylation event leads to a conformational change in ERG that allows ERK to phosphorylate a second serine, S96. S96 phosphorylation then disrupts the interaction between ERG and EZH2/PRC2, allowing ERG to activate gene expression. Together, the interaction of ERG with EWS and the regulation of ERG function by ERK-mediated phosphorylation, represent molecular mechanisms that could serve as targets for therapeutic intervention in ERG-positive prostate cancer.

Citation Format: Vivekananda Kedage, Taylor R. Nicholas, Brady G. Strittmatter, Nagarathinam Selvaraj, Justin A. Budka, Travis J. Jerde, Peter C. Hollenhorst. Regulation of ERG function in prostate cells by phosphorylation and interaction with Ewing’s sarcoma breakpoint protein EWS [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A023.
Abstract A029: Targeting neural transcription factor BRN2 in neuroendocrine prostate tumors
Authors: Munuganti Ravi Shashi Nayana , Thaper Daksh , Zoubeidi Amina
Published: February 2019

Introduction: Resistance to newly developed androgen receptor pathway inhibitors (ARPIs), such as enzalutamide (ENZ), rapidly emerges and patients generally die within two years. In particular, a subset of patients who relapse following ARPI therapy exhibit lineage switching whereby tumors shed their dependence on AR signaling and emerge with neuroendocrine features. These tumors, termed treatment-induced neuroendocrine prostate cancer (t-NEPC), carry an extremely poor prognosis and to date treatment remains decades-old cytotoxic chemotherapy, which carries a short-lived response at the cost of significant toxicity. Thus, the need to develop targeted treatments for this devastating disease is of paramount importance. Recently our group identified the neural transcription factor BRN2 as a major clinically relevant driver of NEPC and aggressive tumor growth, both in vitro and in vivo, suggesting that targeting BRN2 is a promising strategy to prevent neuroendocrine differentiation or treat NEPC.

Methods: Using the integrated power of computational drug discovery platform and biologic testing we identified first-in-field inhibitors for BRN2.

Results: One of the most potent inhibitors identified, Cpd 18, binds to BRN2 using a Drug Affinity Responsive Target Stability (DARTS) assay, decreases the expression of the neuroendocrine genes SOX2, NCAM1, CGA, NSE, PEG10, and N-Myc in BRN2Hi NCI-H660 cells with similar effect in 42DENZR cells. Moreover, Cpd18 displays pronounced antiproliferative activity in NCI-H660 and 42DENZR without any effect on BRN2 low/neg 16DCRPC and LNCaP cells. Combination treatment of Cpd 18 with ENZ prevents the trans-differentiation of 16DCRPC cells into NEPC-like morphology and downregulates the expression of NEPC markers. These outcomes confirmed that the BRN2 is a novel drug target that can help address the problem of ENZ-resistant form of NEPC.

Conclusion: No therapies exist for highly lethal NEPC. Hence, the described work will identify first-in-field inhibitors for BRN2, a central driver of NEPC, and lay the preclinical foundation for the integration of such drugs into the treatment landscape to improve survival and quality of life for patients suffering from deadly form of prostate cancer.

Citation Format: Ravi Shashi Nayana Munuganti, Daksh Thaper, Amina Zoubeidi. Targeting neural transcription factor BRN2 in neuroendocrine prostate tumors [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A029.
Abstract A094: The study of single-nucleotide polymorphisms (SNPs) of tumor suppressor gene in prostate cancer patient of Bangladeshi population
Authors: Imtiaz Hasnain , Afroz Sharmin , Hossain Md. Amir , Bellah Sm Faysal , Rahman Md. Mustafizur
Published: February 2019

Background: Prostate cancer (PCa) is the second leading cause of death among males in present world. The polymorphisms of invasion suppressor gene CDH1 and DNA mismatch repair gene Exo1 have been reported to play a critical role in the tumorigenesis, development, and prognosis of several kinds of cancers, including prostate cancer in some countries. The present study was designed to analyze the contribution of single-nucleotide polymorphisms (SNP) of the CDH1 (-160C/A) and Exo1 (K589E) to prostate cancer susceptibility in Bangladeshi population.

Methods: The study included 100 prostate cancer cases and age-matched 100 healthy controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was used to determine the genetic polymorphisms.

Results: A significant association was found between CDH1 -160 C/A (rs16260) and Exo1 (K589E) polymorphisms and prostate cancer risk. In case of CDH1 -160C/A (rs16260) polymorphism, the frequencies of the three genotypes C/C, C/A, and A/A were 45, 48, and 7% in cases vs. 63, 32, and 5% in controls, respectively. The frequency of C/A heterozygote and combined C/A+A/A genotypes was found to be significantly associated (OR=2.1000, 95% CI= 1.2956. to 4.0905 p=0.013; OR=2.0811, 95% CI= 1.1820-3.6641, p= 0.011, respectively) with increased susceptibility to prostate cancer. The variant A allele also increased the risk of prostate cancer (OR=1.6901, 95% CI= 1.0740 to 2.6597, p= 0.0233). In case of Exo1 (K589E) polymorphism, G/A heterozygote, A/A homozygote, and combined G/A+A/A genotypes were found to be strongly associated (OR= 2.3021, 95% CI=2956. to 4.0905, p=0.0031; OR= 4.8462, 95% CI=1.0198 to 23.0284, p=0.0291; OR= 3.0362, 95% CI=1.7054 to 5.4053, p=0.0001, respectively) with prostate cancer risk. The “A” allele frequency was also found to be significantly associated with increased susceptibility to prostate cancer (OR=2.2955, 95% CI= 1.4529-3.6270, p= 0.0004).

Conclusions: Results of our present study suggest that CDH1 -160 C/A and Exo1 K589E polymorphisms are associated with increased susceptibility to prostate cancer in Bangladeshi population.

Citation Format: Hasnain Imtiaz, Sharmin Afroz, Md. Amir Hossain, Sm Faysal Bellah, Md. Mustafizur Rahman. The study of single-nucleotide polymorphisms (SNPs) of tumor suppressor gene in prostate cancer patient of Bangladeshi population [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A094.
Abstract A097: Regulation of immunophenotype modulation of monocytes-macrophages from M1 to M2 by prostate cancer cell culture supernatant via the transcription factor STAT3
Authors: Solis Raul , Bravo Alejandro , Hernandez Georgina , Bravo Alejandro
Published: February 2019

Background: The transcription factor STAT3 has a prominent innate immunity effect on cancer progression. We determined the regulation of STAT3 in the immunophenotype modulation of macrophages from M1 to M2 induced by the cell culture supernatant of the prostate cancer line PC3.

Methods: Monocyte-macrophages from healthy donors were cultured in the supernatant of PC3 cells, membrane proteins and intracytoplasmic and phosphorylated STAT3 were measured using flow cytometry, and cytokines and growth factors were studied using luminescence. Cytotoxicity and nitric oxide were evaluated via colorimetric assays.

Results: The supernatant of prostate tumor PC3 cells effectively induced macrophages toward an M2 profile, and the expression of phosphorylated STAT3 in the monocytes-macrophages notably increased. In the group of monocytes-macrophages treated with an STAT3 inhibitor, the macrophages were induced towards an M1 phenotype.

Conclusions: In this study, we show that the secretion profile of PC3 prostate cancer cells induces a change in macrophage phenotype from M1 to M2, and this phenomenon is related to phosphorylation of the transcription factor STAT3.

Citation Format: Raul Solis, Alejandro Bravo, Jr., Georgina Hernandez, Alejandro Bravo. Regulation of immunophenotype modulation of monocytes-macrophages from M1 to M2 by prostate cancer cell culture supernatant via the transcription factor STAT3 [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A097.
Abstract B039: Multinucleation precedes the emergence of drug resistance in prostate cancer
Authors: Torga Gonzalo , Lin Ke-Chih , Austin Robert H. , Pienta Kenneth J.
Published: February 2019

Metastatic prostate cancer remains a lethal disease. Taxane chemotherapy used to be restricted to the hormone-resistant metastatic setting in which most patients with castration-resistant prostate cancer (CRPC) develop therapeutic resistance to chemotherapy, achieving limited therapeutic benefit. Recently, two trials demonstrated adjuvant chemohormonal therapy to increase survival in men with high-risk metastatic hormone-sensitive prostate cancer.

Although chemotherapy has been extensively used for decades, how it affects tumor heterogeneity at the molecular level is poorly understood. Experiments with drug gradients in our engineered microfluidic device have shown that docetaxel therapy leads to multinucleation with a stiff distribution pattern of heterogeneous cell populations across the concentration gradient. Furthermore, in these experiments, multinucleation always preceded the development of drug resistance. To confirm these findings, we conducted experiments in conventional cell culture using different chemotherapeutic agents in several prostate cancer cell lines and confirmed multinucleation was a common feature among them. Interestingly, multinucleation occurred to some extent, independently of the mechanism of action of the drug tested.

Aneuploidy, an inherent consequence of multinucleation, has been described as a common characteristic of tumors and proposed to drive tumor progression by increasing the potential for cellular transformation. However, whether this is a cause or a consequence of cancer remains unclear. Aneuploidy can be caused by very diverse molecular mechanisms such as mitotic slippage, endo-replication, or cell fusion. In cancer biology, it has been associated with epithelial-mesenchymal transition as well as with gain of stem-like potential. Recent data in prostate cancer demonstrated the association of multinucleation with chemotherapy, androgen-deprivation therapy, and radiotherapy. Although the data supporting the association of multinucleation with drug resistance are extensive, the effects of these therapies on tumor heterogeneity remain unknown. A better understanding of the resistance mechanisms might reveal new targets for cancer therapy.

In this work, we focus on the characterization of the subset of cells within the tumor population that leads to the multinucleated phenotype, particularly the subpopulation with the potential to reverse polyploidy and repopulate tumor heterogeneity after therapy.

Citation Format: Gonzalo Torga, Ke-Chih Lin, Robert H. Austin, Kenneth J. Pienta. Multinucleation precedes the emergence of drug resistance in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr B039.