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В статье предпринята попытка проанализировать влияние импорта из Китая на российское производство, выделив наиболее значимые аспекты и вытекающие из них последствия. Россия, находящаяся на пути самодостаточности во всех своих сферах, несомненно, должна продолжать идти по данному пути, чтобы минимизировать внешнее влияние.

В статье предпринята попытка проанализировать текущее состояние транспортной системы России, выделив недостатки и указав пути возможных улучшений. Транспортная система страны является важнейшим компонентом ее экономического роста. Модернизированная и эффективная транспортная система может способствовать увеличению товарооборота. Улучшая существующие транспортные системы, предприятия могут сократить время доставки и увеличить объем производства.

The article discusses the approach to the systematization and presentation of heterogeneous information in the textbook. Recommendations are given for the development of teaching materials, taking into account the characteristics of today's "generation Z" students.

The article formulates the specifics of the application of marker symbols for an interactive analogue-digital map. The relevance of the use of marker symbols for an interactive analogue-digital map was substantiated. For the first time, the definition of marker mapping symbols was given.

Currently, data volumes are growing exponentially. Geospatial data is one of the main elements of the concept of Big data. There is a very large number of tools for analyzing Big data, but not all of them take into account the features and have the ability to process geospatial data. The article discusses three popular open analytical tools Hadoop Spatial, GeoSpark, GeoFlink for working with geospatial data of very large volumes. Their architectures, advantages and disadvantages, depending on the execution time and the amount of data used are considered. Processing evaluations were also performed in terms of both streaming and packet data. The experiments were carried out on raster and vector data sets, which are satellite imagery in the visible range, NDVI and NDWI indices, climate indicators (snow cover, precipitation intensity, surface temperature), data from the Open Street Map in the Novosibirsk and Irkutsk Regions.

The article presents a study of the role of interactive map multimedia information for a geoportal. The urgency of the development of interactive map multimedia is shown. The developed technological scheme to create an interactive map with multimedia information is offered.

The article considers the role of multimedia tools and technologies as an integral part of the geomodel used in forecasting emergencies. The efficiency and practicality of using a geomodel integrated with multimedia information is substantiated. A general technological scheme for creating a multimedia geomodel for forecasting emergency situations and its logical structure are proposed.

Background: KRAS activating mutations are reported in 42-54% and inactivation of CDKN2A/p16 in 14-30% of human small intestinal adenocarcinomas, suggesting a role in small intestine adenocarcinogenesis. The phenotypic effects of oncogenic KRAS and p16 loss have not been well characterized in small intestine (SI) adenocarcinogenesis. We targeted LGR5-expressing progenitor cells residing in intestinal crypts for conditional activation of Kras and/or Cdkn2a/p16 deletion to generate a murine model to study the single and combined effects of KRAS and p16 alterations driving gastrointestinal tumorigenesis. We hypothesized that oncogenic Kras and loss of Cdkn2a/p16 drives adenoma-adenocarcinoma sequence in murine SI. Methods: We generated mice with conditional knockout of Cdkn2a/p16 (p16del: IL1bwt/tg;Lgr5-Cre+/−;p16fl/fl;Kraswt/wt), conditional expression of oncogenic KrasG12D (IL1bwt/tg;Lgr5-Cre+/−;KrasG12D/wt), or both alterations (p16del+KrasG12D: IL1bwt/tg;Lgr5-Cre+/−;p16fl/fl;KrasG12D/wt), in mice carrying the human interleukin 1 beta (IL1B) and expressing modified Cre under the Lgr5 promoter. Our control murine model expresses wild type p16 and KRAS (p16wt+Kraswt: IL1bwt/tg;Lgr5-Cre+/−;p16wt/wt;Kraswt/wt). These mice were initially generated to study esophageal adenocarcinogenesis and express the IL1b transgene in esophagus but not in SI. All mice were given bile acid in drinking water daily and were treated with tamoxifen to activate Cre in Lgr5-positive gastrointestinal epithelial progenitor cells. Mice were euthanized and gross and histologic alterations in the SI were examined from 4 to 21 months of age. 3D organoids were isolated from the SI of each mouse model. Results: We examined 89 mice from 4 to 12 months of age. Mice with KRAS activation (KrasG12D) alone infrequently developed SI duodenal adenomas (6.25%: 1/16) compared to 50% (9/18) of mice carrying the combined P16del+KrasG12D genotypes (P=0.005). No adenomas were seen in 27 control and 28 p16del mice. Adenomas with high-grade dysplasia, intramucosal adenocarcinoma, and invasive adenocarcinoma were seen only in older p16del+KrasG12D mice (13-21 months). Single or multiple adenomas and adenocarcinomas were observed only in the duodenal segment of SI and not in the colon. 3D organoids isolated from each mouse model reproduce the intestinal histologic phenotype. Conclusions: Loss of Cdkn2a/p16 and KRASG12D expression synergistically advances adenoma-adenocarcinoma tumorigenesis in the SI duodenum, providing a murine model that recapitulates SI adenocarcinogenesis in vivo. 3D SI organoids provide useful in vitro models for mechanistic studies of SI adenocarcinogenesis. These in vivo and in vitro models will enable identification of key molecular pathways that may be targeted with novel therapies for this rare cancer. Citation Format: Tyler Seckar, Jing Sun, Elena V. Komissarova, Jorge L. Sepulveda, Timothy L. Wang, Antonia R. Sepulveda. Oncogenic KrasG12D and Cdkn2a/p16 knockout in LGR5 expressing progenitor cells synergize to advance adenoma and adenocarcinoma phenotypes in murine small intestine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 24.

Gastric adenocarcinoma (GC) is a leading cause of cancer-related deaths worldwide. The transcription factor gene Friend Leukemia Integration 1 (FLI1) is methylated and downregulated in human GC tissues. Using human GC samples, we determined which cells downregulate FLI1, when FLI1 downregulation occurs, if FLI1 downregulation correlates with clinical-pathologic characteristics, and whether FLI1 plays a role in invasion and/or proliferation of cultured cells. We analyzed stomach tissues from 98 patients [8 normal mucosa, 8 intestinal metaplasia (IM), 7 dysplasia, 91 GC] by immunohistochemistry for FLI1. Epithelial cells from normal, IM, and low-grade dysplasia (LGD) showed strong nuclear FLI1 staining. GC epithelial cells showed significantly less nuclear FLI1 staining as compared to normal epithelium, IM and LGD (P=1.2×10-5, P=1.4×10-6 and P=0.006, respectively). FLI1 expression did not correlate with tumor stage or differentiation, but was associated with patient survival, depending on tumor differentiation. We tested the functional role of FLI1 by assaying proliferation and invasion in cultured GC cells. Lentiviral-transduced FLI1 overexpression in GC AGS cells inhibited invasion by 73.5% (P = 0.001) and proliferation by 31.5% (P = 0.002), as compared to controls. Our results support a combined role for FLI1 as a suppressor of invasiveness and proliferation in gastric adenocarcinoma, specifically in the transition from pre-cancer lesions and dysplasia to invasive adenocarcinoma, and suggest that FLI1 may be a prognostic biomarker of survival in gastric cancers.

The main risk factor for esophageal dysplasia and adenocarcinoma (DAC) is Barrett’s esophagus (BE), characterized by intestinal metaplasia. The critical genomic mechanisms that lead to progression of non-dysplastic BE to DAC remain poorly understood and require analyses of longitudinal patient cohorts and high-resolution assays. We tested BE tissues from 74 patients, including 42 non-progressors from two separate groups of 21 patients each, and 32 progressors (16 in a longitudinal cohort before DAC/pre-progression-BE, and 16 with temporally concurrent but spatially separate DAC/concurrent-BE). We interrogated genome-wide somatic copy number alterations (SCNAs) at the exon level with high-resolution SNP arrays in DNA from formalin-fixed samples histologically confirmed as non-dysplastic BE. The most frequent abnormalities were SCNAs involving FHIT exon 5, CDKN2A/B, or both in 88% longitudinal BE progressors to DAC vs. 24% in both non-progressor groups (P=0.0004). Deletions in other genomic regions were found in 56% of pre-progression-BE but only in 1 non-progressor-BE (P=0.0004). SCNAs involving FHIT exon 5 and CDKN2A/B were also frequently detected in BE temporally concurrent with DAC. TP53 losses were detected in concurrent-BE but not earlier in pre-progression-BE tissues of patients who developed DAC. CDKN2A/p16 immunohistochemistry showed significant loss of expression in BE of progressors vs. non-progressors, supporting the genomic data. Our data suggest a role for CDKN2A/B and FHIT in early progression of BE to dysplasia and adenocarcinoma that warrants future mechanistic research. Alterations in CDKN2A/B and FHIT by high-resolution assays may serve as biomarkers of increased risk of progression to DAC when detected in BE tissues.

Background: Esophageal adenocarcinoma (EAC) develops in pre-cancer columno-glandular hyperplasia, intestinal metaplasia and dysplasia that replaces squamous epithelium and characterizes Barrett’s esophagus (BE). We reported CDKN2A/p16 deletions in up to 69% of BE patients who progressed to dysplasia and EAC, suggesting that p16 loss may drive progression of BE to EAC. LGR5+ cardia like stem cells in the squamocolumnar junction (SCJ) of mouse stomach have been implicated in the cellular origins of BE, and glandular hyperplasia/metaplasia (MGHP) and dysplasia and enhanced inflammation in SCJ were reported in interleukin 1 beta (IL1b) transgenic mice. Our hypothesis is that p16 deletion in LGR5+ cells that give rise to glandular epithelium in SCJ of IL1b mice may accelerate glandular hyperplasia and metaplasia and augment progression to dysplasia and EAC. Design: We tested the hypothesis using conditional knockout of CDKN2A/p16 in IL1b mouse targeting SCJ LGR5+ cells. Model A genotype is p16del: (Lgr5-CreERT2 +/-; IL1b wt/tg; p16 fl/fl) and control model C is p16 wild type (wt): (Lgr5-CreERT2 +/-; IL1b wt/tg; p16 wt/wt). Conditional knockout was induced with intraperitoneal tamoxifen injection. All mice were given bile acid in drinking water, daily. Mice were euthanized and gross and histologic alterations in SCJ, stomach, small intestine (SI) and colon were assessed over time, until 16 months. MGHP was scored as the number of glandular profiles in each section of SCJ, and inflammation and dysplasia were evaluated and scored (0 to 3) in H&E-stained sections. Results: We examined 39 mice: 19 model A (p16del) and 20 model C (p16wt). Mice were grouped into 3 groups from 3 to 16 months. The SCJ of p16del mice exhibited larger areas of hyperplastic cardia-type mucous glands compared to p16wt mice, showing higher numbers of gland profiles (median 17 vs. 8 gland profiles, P=9-5). Higher inflammation scores were also seen in p16del mice ages 3-7 and 7-12 months (P=.036 and P=.013). P16del mice showed higher dysplasia scores overall (P=.013), particularly in mice ages 7-16 months (P=.038). There was a positive correlation between MGHP and inflammation (P=.004) and between inflammation and dysplasia (P=.021). No significant alterations were seen in the stomach, SI, or colon of p16del vs p16wt mice. Conclusions: Loss of p16 alone is not sufficient to develop neoplastic alterations in stomach, SI and colon. Deletion of p16 in LGR5+ cardia-like stem cells in the squamocolumnar junction of mouse stomach, together with IL1b and BA-induced inflammation and cellular injury, is associated with increased MGHP, inflammation, and dysplasia. The model implicates p16 in the early stages of esophageal carcinogenesis, similar to human disease, providing a model to further dissect the contributing molecular mechanisms of progression to esophageal dysplasia and cancer. Citation Format: Jing Sun, Elena V. Komissarova, Caitlin Hills, Tyler Seckar, Gloria H. Su, Jorge Sepulveda, Timothy L. Wang, Antonia R. Sepulveda. CDKN2A/p16 knockout in LGR5+ progenitor cells augments mucous gland hyperplasia/metaplasia, inflammation, and dysplasia in mouse stomach squamocolumnar junction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 929.

Cartographic method of research is widely used in the modern activities of the Ministry of Emergency Situations of Russia. Introduction of GIS products and use of digital and electronic maps allows automating the process of creating graphic documents. A number of unsolved problems remain: an outdated system of symbols, old methods of geoinformation mapping. The article discusses problems that arise when using the existing system of conventional symbols for mapping command posts, analyzes the existing regulatory documents. Causes of such problems are identified and possible solutions are offened.

Despite a good enough degree so traditional printed maps, however you can still find new ways to increase information card capacity and thus partially resolve the existing contradiction between the clarity and readability of the maps. One such way is to use QR code technology. The article describes the basic features of QR codes, to ensure that the interest in this technology. The research identified a number of advantages of using QR codes when creating traditional cards. Technology research with the use of QR codes in cartography has both scientific and practical value in terms of creating traditional cards for various content and destination.

Presently there is no general method of creating and using traditional (paper) maps using mobile devices and QR codes. The article formulates the purpose and objectives of the research in the development of a new kind of interactive cartographic work. As a result of studying the features of QR codes, considering the possibilities of expanding the information capacity of traditional printed maps using mobile devices and QR codes, the basic possibilities of using them when creating a new type of map, advantages and disadvantages of using mobile devices and QR codes when creating and using a traditional paper map are highlighted.

This report discusses the problem of wide axial ratio beamwidth and high front-to-back ratio compact antenna design problem. As a prospective solution, a system design, employing crossed dipole element and circular reflector with a curved sidewall, is proposed. In the first step of research, the reflector’s influence on radiation characteristics is examined using a simplified model. Numerical analysis shows the potential front-to-back ratio value around 36 dB while the farfield radiation pattern in the upper hemisphere is close to axisymmetric and has a low cross-polarization component level for the same reflector’s geometry.

The results of a constructively simple high-tech small-sized integrated element of the Ka-band electric beam scanning reflective phased array antenna (PAA) development are shown. Beam scanning sector of the phased array antenna is up to 60 by both coordinates. The PAA element is based on the Faraday type waveguide ferrite phase shifter, which works on the circular polarized electromagnetic waves. More than 30000 PAA elements has been produced, 100% control and statistical data processing of their main characteristics has been carried out. Graph for initial phases and steepness of linearized phase characteristics, medium and maximum losses of PAA elements are performed. Average value of the average deviation for parameters which is important for PAA and the beam control system design have been determined. The developed element can be used in PAA which have different sizes, forms and sectors of electrical beam scanning. Small deviations in the characteristics of PAA elements from the linearized characteristics indicate the high quality of the production and their identity, which makes it possible to create PAA without taking into account the real individual parameters of mass-produced PAA elements.

The electrodynamic analysis and calculation of the phasing structure of a waveguide ferrite Faraday phase shifter with longitudinal magnetization of a ferrite rod with a square cross-section are performed. In the strict electrodynamic formulation, the Galerkin method solves the key problem for the design of the phase shifter: the problem of eigenwaves finding with a second-order differential magnetic operator in the projection procedure for a rectangular waveguide with a transversely inhomogeneous ferrite-dielectric filling under longitudinal magnetization. The problem is reduced to solving the complete eigenvalue problem of a complex matrix. The computational algorithm, implemented as a computer program in DELPHI, gives in one procedure the numerical values of the coefficients of the decomposition of fields in the form of eigenvectors of the matrix, and the coefficients of wave propagation in the form of its eigenvalues. The results of calculations of the phasing structure activity for two particular cases of its implementation are presented: on the basis of a ferrite rod both with a conductive coating and without a coating, it is shown that in the second case the activity of the phase shifter is 1.5 times higher. For the real parameters of the ferrite medium, the length of the phasing structure is calculated depending on the transverse dimensions of the ferrite rod and the square waveguide, which ensures the creation of a 360° controlled phase shift.

Gastric adenocarcinoma (GC) is a leading cause of cancer-related deaths worldwide. The transcription factor gene Friend Leukemia Integration 1 (FLI1) is methylated and downregulated in human GC tissues. Using human GC samples, we determined which cells downregulate FLI1, when FLI1 downregulation occurs, if FLI1 downregulation correlates with clinical-pathologic characteristics, and whether FLI1 plays a role in invasion and/or proliferation of cultured cells. We analyzed stomach tissues from 98 patients [8 normal mucosa, 8 intestinal metaplasia (IM), 7 dysplasia, 91 GC] by immunohistochemistry for FLI1. Epithelial cells from normal, IM, and low-grade dysplasia (LGD) showed strong nuclear FLI1 staining. GC epithelial cells showed significantly less nuclear FLI1 staining as compared to normal epithelium, IM and LGD (P=1.2×10-5, P=1.4×10-6 and P=0.006, respectively). FLI1 expression did not correlate with tumor stage or differentiation, but was associated with patient survival, depending on tumor differentiation. We tested the functional role of FLI1 by assaying proliferation and invasion in cultured GC cells. Lentiviral-transduced FLI1 overexpression in GC AGS cells inhibited invasion by 73.5% (P = 0.001) and proliferation by 31.5% (P = 0.002), as compared to controls. Our results support a combined role for FLI1 as a suppressor of invasiveness and proliferation in gastric adenocarcinoma, specifically in the transition from pre-cancer lesions and dysplasia to invasive adenocarcinoma, and suggest that FLI1 may be a prognostic biomarker of survival in gastric cancers.

microRNAs (miRNAs) play key roles in cancer, but their propensity to couple their targets as competing endogenous RNAs (ceRNAs) has only recently emerged. Multiple models have studied ceRNA regulation, but these models did not account for the effects of co-regulation by miRNAs with many targets. We modeled ceRNA and simulated its effects using established parameters for miRNA/mRNA interaction kinetics while accounting for co-regulation by multiple miRNAs with many targets. Our simulations suggested that co-regulation by many miRNA species is more likely to produce physiologically relevant context-independent couplings. To test this, we studied the overlap of inferred ceRNA networks from four tumor contexts—our proposed pan-cancer ceRNA interactome (PCI). PCI was composed of interactions between genes that were co-regulated by nearly three-times as many miRNAs as other inferred ceRNA interactions. Evidence from expression-profiling datasets suggested that PCI interactions are predictive of gene expression in 12 independent tumor- and non-tumor contexts. Biochemical assays confirmed ceRNA couplings for two PCI subnetworks, including oncogenes CCND1, HIF1A and HMGA2, and tumor suppressors PTEN, RB1 and TP53. Our results suggest that PCI is enriched for context-independent interactions that are coupled by many miRNA species and are more likely to be context independent.

The diverse roles of protein kinase C-δ (PKCδ) in cellular growth, survival, and injury have been attributed to stimulus-specific differences in PKCδ signaling responses. PKCδ exerts membrane-delimited actions in cells activated by agonists that stimulate phosphoinositide hydrolysis. PKCδ is released from membranes as a Tyr(313)-phosphorylated enzyme that displays a high level of lipid-independent activity and altered substrate specificity during oxidative stress. This study identifies an interaction between PKCδ's Tyr(313)-phosphorylated hinge region and its phosphotyrosine-binding C2 domain that controls PKCδ's enzymology indirectly by decreasing phosphorylation in the kinase domain ATP-positioning loop at Ser(359). We show that wild-type (WT) PKCδ displays a strong preference for substrates with serine as the phosphoacceptor residue at the active site when it harbors phosphomimetic or bulky substitutions at Ser(359.) In contrast, PKCδ-S359A displays lipid-independent activity toward substrates with either a serine or threonine as the phosphoacceptor residue. Additional studies in cardiomyocytes show that oxidative stress decreases Ser(359) phosphorylation on native PKCδ and that PKCδ-S359A overexpression increases basal levels of phosphorylation on substrates with both phosphoacceptor site serine and threonine residues. Collectively, these studies identify a C2 domain-pTyr(313) docking interaction that controls ATP-positioning loop phosphorylation as a novel, dynamically regulated, and physiologically relevant structural determinant of PKCδ catalytic activity.

We introduce and validate a new precision oncology framework for the systematic prioritization of drugs targeting mechanistic tumor dependencies in individual patients. Compounds are prioritized on the basis of their ability to invert the concerted activity of master regulator proteins that mechanistically regulate tumor cell state, as assessed from systematic drug perturbation assays. We validated the approach on a cohort of 212 gastroenteropancreatic neuroendocrine tumors (GEP-NETs), a rare malignancy originating in the pancreas and gastrointestinal tract. The analysis identified several master regulator proteins, including key regulators of neuroendocrine lineage progenitor state and immunoevasion, whose role as critical tumor dependencies was experimentally confirmed. Transcriptome analysis of GEP-NET-derived cells, perturbed with a library of 107 compounds, identified the HDAC class I inhibitor entinostat as a potent inhibitor of master regulator activity for 42% of metastatic GEP-NET patients, abrogating tumor growth in vivo. This approach may thus complement current efforts in precision oncology.

Activation of TLR by ssRNA after FcγR-mediated phagocytosis of immune complexes (IC) may be relevant in autoimmune-associated congenital heart block (CHB) where the obligate factor is a maternal anti-SSA/Ro Ab and the fetal factors, protein/RNA on an apoptotic cardiocyte and infiltrating macrophages. This study addressed the hypothesis that Ro60-associated ssRNAs link macrophage activation to fibrosis via TLR engagement. Both macrophage transfection with noncoding ssRNA that bind Ro60 and an IC generated by incubation of Ro60-ssRNA with an IgG fraction from a CHB mother or affinity purified anti-Ro60 significantly increased TNF-α secretion, an effect not observed using control RNAs or normal IgG. Dependence on TLR was supported by the significant inhibition of TNF-α release by IRS661 and chloroquine. The requirement for FcγRIIIa-mediated delivery was provided by inhibition with an anti-CD16a Ab. Fibrosis markers were noticeably increased in fetal cardiac fibroblasts after incubation with supernatants generated from macrophages transfected with ssRNA or incubated with the IC. Supernatants generated from macrophages with ssRNA in the presence of IRS661 or chloroquine did not cause fibrosis. In a CHB heart, but not a healthy heart, TLR7 immunostaining was localized to a region near the atrioventricular groove at a site enriched in mononuclear cells and fibrosis. These data support a novel injury model in CHB, whereby endogenous ligand, Ro60-associated ssRNA, forges a nexus between TLR ligation and fibrosis instigated by binding of anti-Ro Abs to the target protein likely accessible via apoptosis.

Polo-like kinase 3 (Plk3) is an important mediator of the cellular responses to genotoxic stresses. In this study, we examined the physiologic function of Plk3 by generating Plk3-deficient mice. Plk3−/− mice displayed an increase in weight and developed tumors in various organs at advanced age. Many tumors in Plk3−/− mice were large in size, exhibiting enhanced angiogenesis. Plk3−/− mouse embryonic fibroblasts were hypersensitive to the induction of hypoxia-inducible factor-1α (HIF-1α) under hypoxic conditions or by nickel and cobalt ion treatments. Ectopic expression of the Plk3-kinase domain (Plk3-KD), but not its Polo-box domain or a Plk3-KD mutant, suppressed the nuclear accumulation of HIF-1α induced by nickel or cobalt ions. Moreover, hypoxia-induced HIF-1α expression was tightly associated with a significant down-regulation of Plk3 expression in HeLa cells. Given the importance of HIF-1α in mediating the activation of the “survival machinery” in cancer cells, these studies strongly suggest that enhanced tumorigenesis in Plk3-null mice is at least partially mediated by a deregulated HIF-1 pathway. [Cancer Res 2008;68(11):4077–85]

Drinking arsenic-contaminated water is associated with increased risk of neoplasias of the skin, lung, bladder and possibly other sites, as well as other diseases. Earlier, we showed that human lymphoblast lines from different normal unexposed donors showed variable sensitivities to the toxic effects of arsenite. In the present study, we used microarray analysis to compare the basal gene expression profiles between two arsenite-resistant (GM02707, GM00893) and two arsenite-sensitive lymphoblast lines (GM00546, GM00607). A number of genes were differentially expressed in arsenite-sensitive and arsenite-resistant cells. Among these, gamma-glutamyltranspeptidase 1 (GGT1) and NF kappa B inhibitor-epsilon (NFKBIE) showed higher expression levels in arsenite-resistant cells. RT-PCR analysis with gene-specific primers confirmed these results. Reduction of GGT1 expression level in arsenite-resistant lymphoblasts with GGT1-specific siRNA resulted in increased cell sensitivity to arsenite. In conclusion, we have demonstrated for the first time that expression levels of GGT1 and possibly NFKBIE might be useful as biomarkers of genetic susceptibility to arsenite. Expression microarrays can thus be exploited for identifying additional biomarkers of susceptibility to arsenite and to other toxicants.

Gastric cancers are the most frequent gastric malignancy and usually arise in the sequence of Helicobacter pylori-associated chronic gastritis. CpG methylation is a central mechanism of epigenetic gene regulation affecting cancer-related genes, and occurs early in gastric carcinogenesis. DNA samples from non-metaplastic gastric mucosa with variable levels of gastritis (non-metaplastic mucosa), intestinal metaplasia, or gastric cancer were screened with methylation arrays for CpG methylation of cancer-related genes and 30 gene targets were further characterized by high-definition bisulfite next-generation sequencing. In addition, data from The Cancer Genome Atlas were analyzed for correlation of methylation with gene expression. Overall, 13 genes had significantly increased CpG methylation in gastric cancer vs non-metaplastic mucosa (BRINP1, CDH11, CHFR, EPHA5, EPHA7, FGF2, FLI1, GALR1, HS3ST2, PDGFRA, SEZ6L, SGCE, and SNRPN). Further, most of these genes had corresponding reduced expression levels in gastric cancer compared with intestinal metaplasia, including novel hypermethylated genes in gastric cancer (FLI1, GALR1, SGCE, and SNRPN), suggesting that they may regulate neoplastic transformation from non-malignant intestinal metaplasia to cancer. Our data suggest a tumor-suppressor role for FLI1 in gastric cancer, consistent with recently reported data in breast cancer. For the genes with strongest methylation/expression correlation, namely FLI1, the expression was lowest in microsatellite-unstable tumors compared with other gastric cancer molecular subtypes. Importantly, reduced expression of hypermethylated BRINP1 and SGCE was significantly associated with favorable survival in gastric cancer. In summary, we report novel methylation gene targets that may have functional roles in discrete stages of gastric carcinogenesis and may serve as biomarkers for diagnosis and prognosis of gastric cancer.

Arsenite is the most likely carcinogenic form of arsenic in the environment. Previously, expression cloning for cDNAs whose overexpression confers arsenite-resistance in Chinese hamster V79 cells identified two genes: fau and a novel gene, asr2. The fau gene encodes a ubiquitin-like protein (here called FUBI) fused to the ribosomal S30 protein. Since the expression of the fox sequence (antisense to fau) increased the tumorigenicity of a mouse sarcoma virus, it was proposed that fau might be a tumor suppressor gene. We intended to test its ability to block arsenite-induced transformation of human osteogenic sarcoma (HOS) cells to anchorage-independence. Instead, we found that overexpressing fau itself was able to transform HOS cells. When the two domains were expressed separately, only FUBI was transforming and only the S30 domain conferred arsenite resistance. An incidental finding was the transforming activity of the selectable marker, hyg. FUBI belongs to the ubiquitin-like protein group that is capable of forming conjugates to other proteins, although none have so far been identified. Alternatively, FUBI may act as a substitute or inhibitor of ubiquitin, to which it is most closely related, or to close ubiquitin-like relatives UCRP, FAT10, and/or Nedd8.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most common cancers in the United States. The five-year survival rate for patients with PDAC remains dismal. Identification of biomarkers for early diagnosis of PDAC and pre-cancer pancreatic intraepithelial neoplasia (PanIN) lesions with risk of progression to PDAC is critically needed. We hypothesized that differentially expressed genes and regulatory pathways in PanIN and PDAC compared to normal duct epithelium (ND) may represent biomarkers of development of malignancy. We used Affymetrix Human Transcriptome Arrays 2.0 to establish gene expressionprofiles in ND, low-grade PanIN, and PDAC epithelium. Total RNA was isolated after laser capture microdissection (LCM) of frozen tissue sections and then used for producing hybridization-ready DNA. Hybridization quality control was performed with Expression Console 1.4 software and background corrected/normalized data were analyzed with Transcriptome Analysis Console (TAC) 3.1 and the ASSIGN algorithm. We tested RNA from 22 LCM samples (9 PDACs, 5 PanINs, and 8 ND), including 4 matched trios of ND, PanIN and PDAC from the same patients. Differential expression analysis with one-way between subject ANOVA revealed over 2000 genes differentially expressed in PDAC and PanIN vs. ND group (filter criteria up/down >2; ANOVA p1.5; ANOVA p Citation Format: Elena V. Komissarova, Jorge Sepulveda, Sarawut Kongkarnka, Maryam Shirazi, Brynn Levy, Claudia Cujar, Antonia R. Sepulveda. Human transcriptome alterations in pre-cancer and cancer epithelium identify candidate biomarkers of progression to pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 714. doi:10.1158/1538-7445.AM2017-714

Gastric adenocarcinoma (GC) is the 5th most common cancer worldwide but is the 3rd leading cause of cancer death. FLI1 (Friend leukemia integration-1) is an ETS family transcription factor that regulates genes involved in proliferation and differentiation. FLI1 is implicated in tumorigenesis, such as in Ewing’s sarcoma where a translocation creates an EWS-FLI1 oncogenic fusion protein. However, few studies have examined the role of FLI1 in carcinomas. In human breast cancer, overexpression of FLI1 led to inhibition of apoptosis, thereby promoting survival and malignant potential. In functional studies in a murine breast cancer model, however, downregulation of FLI1 increased malignant potential. In human GCs, we recently reported that FLI1 expression is inversely correlated with its promoter CpG methylation of the FLI1. To determine if decreased expression occurs in GC epithelial cells or background non-epithelial cells, we analyzed 91 human GC tumors by immunohistochemistry (IHC) for FLI1 and compared them to normal gastric mucosa and intestinal metaplasia (IM) using an IHC composite scoring system accounting for intensity and percentage of epithelial cells expressing FLI1. We found that FLI1 is strongly expressed in normal gastric glandular epithelium and in IM, and that decreased expression was seen in most human GCs (P < 1x10-17 vs normal, P < 1x10-26 vs IM). These findings suggest that FLI1 acts as a tumor suppressor gene in GC. To test this hypothesis, we used an invasion assay and the human GC cell lines NUGC3 and SNU638, which have little to no FLI1 expression, respectively. Cultured GC cells were transduced to overexpress FLI1 or control, along with an eGFP reporter from an IRES (Lv203, Genecopoeia). After selection by puromycin, these GC cells were plated in serum-free media in the upper chamber on a Matrigel coated 8µm pore opaque membrane. Complete media with 10% fetal calf serum was plated in the lower chamber. Images were obtained of the lower membrane with an inverted fluorescent microscope and cellSens imaging software. Overexpression of FLI1 significantly decreased invasion by NUGC3 cells at 24 hours (P = 0.013) but not at 48 hours (P = 0.268) as compared to control. Overexpression of FLI1 significantly decreased invasion by SNU638 cells at both 24 and 48 hours (P = 0.027 and 0.012, respectively) as compared to control. Since NUGC3 cells have low FLI1 expression, we knocked down FLI1 by using a FLI1 shRNA lentiviral system with mCherry from an IRES as a reporter (LvRU6MP, Genecopoeia). Using the same invasion assay, knockdown of FLI1 trended towards a significant increase in invasion as compared to control at 48 hrs (P = 0.14), but not at 24 hrs (P = 0.73). In summary, the combined observations in human GC tissue samples and the functional analyses in GC cells support a tumor suppressor role for FLI1 in human GC and also suggest that FLI1 and/or its target genes may be involved in regulatory mechanisms driving invasive properties of GC. Citation Format: Armando Del Portillo, Elena V. Komissarova, Anne Koehne de Gonzalez, Aqiba Bokhari, Helen Remotti, Jorge Sepulveda, Antonia Sepulveda. Functional role of Friend Leukemia Integration-1 (FLI1) in gastric carcinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5532. doi:10.1158/1538-7445.AM2017-5532

Rationale : Binding of maternal anti-Ro/La antibodies to cognate antigen expressed on apoptotic cardiocytes decreases clearance by healthy cardiocytes, which may contribute to the development of autoimmune associated congenital heart block and fatal cardiomyopathy. Objective : Given recent evidence implicating the urokinase plasminogen activator receptor (uPAR) as a “don't eat me” signal during efferocytosis, experiments addressed whether surface bound anti-Ro antibodies inhibit apoptotic cell removal via an effect on the expression/function of the urokinase-type plasminogen activator protease uPA/uPAR system. Methods and Results : As assessed by flow cytometry and confocal microscopy, uPAR colocalizes and interacts with Ro60 on the surface of apoptotic human fetal cardiocytes. Blocking of uPAR enhances phagocytosis of apoptotic cardiocytes by healthy cardiocytes and reverses the anti-Ro60–dependent impaired clearance of apoptotic cardiocytes. Binding of anti-Ro60 antibodies to apoptotic cardiocytes results in increased uPAR expression, as well as enhanced uPA activity. The binding of anti-Ro60 did not alter other surface molecules involved in cell recognition (calreticulin, CD31, or CD47). Conclusions : These data suggest that increased uPAR expression and uPA activity induced by anti-Ro60 binding to the apoptotic fetal cardiocyte provide a molecular basis by which these antibodies inhibit efferocytosis and ultimately lead to scar of the fetal conduction system and working myocardium.

The recurrence rate of anti-SSA/Ro-associated congenital heart block (CHB) is 17%. Sustained reversal of third-degree block has never been achieved. Based on potential reduction of maternal autoantibody titers as well as fetal inflammatory responses, intravenous immunoglobulin (IVIG) was evaluated as preventive therapy for CHB.A multicenter, prospective, open-label study based on Simon's 2-stage optimal design was initiated. Enrollment criteria included the presence of anti-SSA/Ro antibodies in the mother, birth of a previous child with CHB/neonatal lupus rash, current treatment withor = 20 mg/day of prednisone, and12 weeks pregnant. IVIG (400 mg/kg) was given every 3 weeks from week 12 to week 24 of gestation. The primary outcome was the development of second-degree or third-degree CHB.Twenty mothers completed the IVIG protocol before the predetermined stopping rule of 3 cases of advanced CHB in the study was reached. CHB was detected at 19, 20, and 25 weeks; none of the cases occurred following the finding of an abnormal PR interval on fetal Doppler monitoring. One of these mothers had 2 previous children with CHB. One child without CHB developed a transient rash consistent with neonatal lupus. Sixteen children had no manifestations of neonatal lupus at birth. No significant changes in maternal titers of antibody to SSA/Ro, SSB/La, or Ro 52 kd were detected over the course of therapy or at delivery. There were no safety issues.This study establishes the safety of IVIG and the feasibility of recruiting pregnant women who have previously had a child with CHB. However, IVIG at low doses consistent with replacement does not prevent the recurrence of CHB or reduce maternal antibody titers.

The percentages of cells with different types of cell surface relief were determined in cell suspensions derived from monolayer cultures. Primary cultures of rat embryo fibroblasts (REF) and cell lines REF (LT) and REF-1, immortalized cells of which preserved normal phenotypic characteristics of the initial primary culture REF, as well as morphologically transformed tumorigenic lines REF (LT) ras and REF-2EJ were studied. In REF suspensions the cells with the blebbed type of surface relief were shown to be predominant as compared with those with microvillus relief whereas cell suspensions derived from both immortalized and fully transformed cultures display the reverse ratio of cells with those types of surface relief. Therefore, the pattern of cell surface relief in cell suspensions derived from fibroblastic monolayer cultures may serve as a morphological marker of the initial stage of neoplastic transformation-immortalization when typical morphological signs of cell transformation are not yet manifested in monolayer cultures.

Arsenite is an environmental pollutant. Exposure to inorganic arsenic in drinking water is associated with elevated cancer risk, especially in skin. Arsenite alone does not cause skin cancer in animals, but arsenite can enhance the carcinogenicity of solar UV. Arsenite is not a significant mutagen at non-toxic concentrations, but it enhances the mutagenicity of other carcinogens. The tumor suppressor protein P53 and nuclear enzyme PARP-1 are both key players in DNA damage response. This laboratory demonstrated earlier that in cells treated with arsenite, the P53-dependent increase in p21WAF1/CIP1 expression, normally a block to cell cycle progression after DNA damage, is deficient. Here we show that although long-term exposure of human keratinocytes (HaCaT) to a nontoxic concentration (0.1 μM) of arsenite decreases the level of global protein poly(ADP-ribosyl)ation, it increases poly(ADP-ribosyl)ation of P53 protein and PARP-1 protein abundance. We also demonstrate that exposure to 0.1 μM arsenite depresses the constitutive expression of p21 mRNA and P21 protein in HaCaT cells. Poly(ADP-ribosyl)ation of P53 is reported to block its activation, DNA binding and its functioning as a transcription factor. Our results suggest that arsenite's interference with activation of P53 via poly(ADP-ribosyl)ation may play a role in the comutagenic and cocarcinogenic effects of arsenite.

Objectives To understand if there exists a functional interaction between arsenic trioxide and paclitaxel in vitro. Materials and methods HeLa and HCT116 (rho53(+/+) and rho53(-/-)) cells were treated with As2O3 and/or paclitaxel for various times. Treated cells were collected for analyses using a combination of flow cytometry, fluorescence microscopy and Western blotting. Results Because As(2)O(3) is capable of inhibiting tubulin polymerization and inducing mitotic arrest, we examined whether there existed any functional interaction between As(2)O(3) and paclitaxel, a well-known microtubule poison. Flow cytometry and fluorescence microscopy revealed that although As(2)O(3) alone caused a moderate level of mitotic arrest, it greatly attenuated paclitaxel-induced mitotic arrest in cells with p53 deficiency. Western blot analysis showed that As(2)O(3) significantly blocked phosphorylation of BubR1, Cdc20, and Cdc27 in cells treated with paclitaxel, suggesting that arsenic compromised the activation of the spindle checkpoint. Our further studies revealed that the attenuation of paclitaxel-induced mitotic arrest by As(2)O(3) resulted primarily from sluggish cell cycle progression at S phase but not enhanced mitotic exit. Conclusion The observations that As(2)O(3) has a negative impact on the cell cycle checkpoint activation by taxol should have significant clinical implications because the efficacy of taxol in the clinics is associated with its ability to induce mitotic arrest and subsequent mitotic catastrophe.

Arsenic is a common environmental contaminant of our air, water and food, but not every individual who drinks arsenic-contaminated water shows clinical signs of toxicity. Large inter-individual variations are also found in arsenite-induced aneuploidy, chromosome aberrations and sister chromatid exchanges in peripheral blood lymphocytes from different human donors. Lymphoblasts are virally immortalized lymphocytes that retain most of the properties of lymphocytes. Individual lymphoblast cell lines retained their arsenite sensitivity after cryopreservation and subsequent revival. We measured the accumulation of 73[As]-arsenite into lymphoblast lines derived from 11 normal individuals. Arsenite accumulation rate varied 6.3 fold between the slowest and the fastest subjects. Assays in 14 lymphoblast lines showed variability to the toxic effects of arsenite, as measured by growth inhibition. Lymphoblast lines also vary with regard to their growth rates, but there is no relationship between growth rate and arsenite sensitivity. Surprisingly, we also found no correlation between arsenite accumulation rate and cellular sensitivity to growth inhibition, suggesting that the arsenite accumulation rate may not be the main determinant of cellular sensitivity to arsenic. We were also unable to detect evidence for a human homolog for the yeast arsenite efflux gene ACR3, using RT-PCR.

DNA damage is preferentially repaired in the transcribed strand of many active genes. Although the concept of DNA repair coupled with transcription has been widely accepted, its mechanisms remain elusive. We recently reported that in Chinese hamster ovary cells while ultraviolet light-induced cyclobutane pyrimidine dimers (CPDs) are preferentially repaired in the transcribed strand of dihydrofolate reductase gene, CPDs are efficiently repaired in both strands of adenine phosphoribosyltransferase (APRT) locus, in either a transcribed or nontranscribed APRT gene (1). These results suggested that the transcription dependence of repair may depend on genomic context. To test this hypothesis, we constructed transfectant cell lines containing a single, actively transcribed APRT gene, integrated at different genomic sites. Mapping of CPD repair in the integrated APRT genes in three transfectant cell lines revealed two distinct repair patterns, either preferential repair of CPDs in the transcribed strand or very poor repair in both strands. Similar kinetics of micrococcal nuclease digestion were seen for all three transfectant APRT gene domains and endogenous APRT locus. Our results suggest that both the efficiency and strand-specificity of repair of an actively transcribed gene are profoundly affected by genomic context but do not reflect changes in first order nucleosomal structure.

Chromosomal analysis of immortalized rat embryo fibroblasts (IE5) and transformed by HPV18 E6+E7 genes (A4E5) or HPV16 E7 alone (trB4; trF8; trC2) variants have been done. Transformed cell lines represented heterogeneous cell populations containing neardiploid subpopulations with 41-44 chromosomes and also heterogeneous polyploid cells in contrast to immortal cells IE5 that contained normal number of chromosomes-42. In transformed cells the abnormalities of interphase nuclei (giant-, micro-, apoptotic nuclei) were observed which could reflect genomic instability of polyploid cells. Several chromosomal alterations were revealed in immortal IE5 cells, but only reciprocal translocation t(8; 10) (q22q12.3) was stable and kept in cells transformed by HPV18 E6+E7 genes or HPV16 E7 alone. We can conclude that genomic instability and clonal expansion of the cells with specific chromosomal alterations contribute to HPV-mediated transformation.

Loss of heterozygosity (LOH) in chromosome 6 in human squamous cervical carcinomas was analyzed in the long and short arms of the chromosome using 3 pairs of primers each. In all cases, normal adjacent tissue was used as control. Among 51 cases analyzed, we identified LOH or microsatellite instability in 23% using primer D6S291 (located at position 6p21.3) and in 11% using primers D6S308 (6q16.3–6q27) and D6S270 (6q22.3–6q23.2). On the contrary, no significant LOH or genomic instabilities were detected with primers D6S306 (6p22.3–6p21.2), D6S299 (6p22.3–6p21.3) and D6S287 (6q21–6q23.3). Our results thus suggest the existence of instable loci at 3 regions of chromosome 6. Whether these loci contain putative tumor-suppressor genes or genes involved in cell cycle control remains unknown. © 1996 Wiley-Liss, Inc.

Organs of mice with leukemia induced by Rauscher and Mazurenko viruses contain different amounts of virus-specific RNA. In Mazurenko virus-induced leukemia, virus-specific RNA content reached the maximum in the thymus and lymph nodes. Hybridization level of RLV 3H DNA transcript with RNA from mouse cells with RLV-induced leukemia amounted to 90% while in cells of mice with Mazurenko virus-induced leukemia it did not exceed 60%.