Yukyung Jun

miRGator v2.0 : an integrated system for functional investigation of microRNAs

miRGator is an integrated database of microRNA (miRNA)-associated gene expression, target prediction, disease association and genomic annotation, which aims to facilitate functional investigation of miRNAs. The recent version of miRGator v2.0 contains information about (i) human miRNA expression profiles under various experimental conditions, (ii) paired expression profiles of both mRNAs and miRNAs, (iii) gene expression profiles under miRNA-perturbation (e.g. miRNA knockout and overexpression), (iv) known/predicted miRNA targets and (v) miRNA-disease associations. In total, >8000 miRNA expression profiles, ∼300 miRNA-perturbed gene expression profiles and ∼2000 mRNA expression profiles are compiled with manually curated annotations on disease, tissue type and perturbation. By integrating these data sets, a series of novel associations (miRNA–miRNA, miRNA–disease and miRNA–target) is extracted via shared features. For example, differentially expressed genes (DEGs) after miRNA knockout were systematically compared against miRNA targets. Likewise, differentially expressed miRNAs (DEmiRs) were compared with disease-associated miRNAs. Additionally, miRNA expression and disease-phenotype profiles revealed miRNA pairs whose expression was regulated in parallel in various experimental and disease conditions. Complex associations are readily accessible using an interactive network visualization interface. The miRGator v2.0 serves as a reference database to investigate miRNA expression and function (http://miRGator.kobic.re.kr).

Rapid emergence and mechanisms of resistance by U87 glioblastoma cells to doxorubicin in an in vitro tumor microfluidic ecology

Significance Drug resistance is among the most critical problems in cancer treatment. This proof-of-principle study demonstrates that a combination of in vitro microecology and deep sequencing could provide an extremely efficient method to elucidate the in vivo resistance mechanisms of cancer drugs and predict if drug resistance is likely to occur. Remarkably, drug resistance was seen within 7 d in our microfluidic chip, enabling characterization of molecular mechanisms within a month. This would be of great value for clinicians in selecting drugs likely to be slow in drug resistance emergence, and choose therapies for resistant tumors more effectively. In principle, our microecology technology can be used for many combinations of cancer types and drugs. In vitro prediction of the probable rapid emergence of resistance to a drug in tumors could act to winnow out potential candidates for further costly development. We have developed a microfluidic device consisting of ∼500 hexagonal microcompartments that provides a complex ecology with wide ranges of drug and nutrient gradients and local populations. This ecology of a fragmented metapopulation induced the drug resistance in stage IV U87 glioblastoma cells to doxorubicin in 7 d. Exome and transcriptome sequencing of the resistant cells identified mutations and differentially expressed genes. Gene ontology and pathway analyses of the genes identified showed that they were functionally relevant to the established mechanisms of doxorubicin action. Specifically, we identified (i) a frame-shift insertion in the filamin-A gene, which regulates the influx and efflux of topoisomerase II poisons; (ii) the overexpression of aldo-keto reductase enzymes, which convert doxorubicin into doxorubicinol; and (iii) activation of NF-κB via alterations in the nucleotide-binding oligomerization domain (NOD)-like receptor signaling pathway from mutations in three genes (CARD6, NSD1, and NLRP13) and the overexpression of inflammatory cytokines. Functional experiments support the in silico analyses and, together, demonstrate the effects of these genetic changes. Our findings suggest that, given the rapid evolution of resistance and the focused response, this technology could act as a rapid screening modality for genetic aberrations leading to resistance to chemotherapy as well as counter selection of drugs unlikely to be successful ultimately.

Interaction of tankyrase and peroxiredoxin II is indispensable for the survival of colorectal cancer cells

Mammalian 2-Cys peroxiredoxin (Prx) enzymes are overexpressed in most cancer tissues, but their specific signaling role in cancer progression is poorly understood. Here we demonstrate that Prx type II (PrxII) plays a tumor-promoting role in colorectal cancer by interacting with a poly(ADP-ribose) polymerase (PARP) tankyrase. PrxII deletion in mice with inactivating mutation of adenomatous polyposis coli (APC) gene reduces intestinal adenomatous polyposis via Axin/β-catenin axis and thereby promotes survival. In human colorectal cancer cells with APC mutations, PrxII depletion consistently reduces the β-catenin levels and the expression of β-catenin target genes. Essentially, PrxII depletion hampers the PARP-dependent Axin1 degradation through tankyrase inactivation. Direct binding of PrxII to tankyrase ARC4/5 domains seems to be crucial for protecting tankyrase from oxidative inactivation. Furthermore, a chemical compound targeting PrxII inhibits the expansion of APC-mutant colorectal cancer cells in vitro and in vivo tumor xenografts. Collectively, this study reveals a redox mechanism for regulating tankyrase activity and implicates PrxII as a targetable antioxidant enzyme in APC-mutation-positive colorectal cancer.2-Cys peroxiredoxin (Prx) enzymes are highly expressed in most cancers but how they promote cancer progression is unclear. Here the authors show that in colorectal cancers with APC mutation, PrxII binds to tankyrase and prevents its oxidative inactivation, thereby preventing Axin1-dependent degradation of ²b-catenin.

Characterization of SLC22A18 as a tumor suppressor and novel biomarker in colorectal cancer

SLC22A18, solute carrier family 22, member 18, has been proposed to function as a tumor suppressor based on its chromosomal location at 11p15.5, mutations and aberrant splicing in several types of cancer and down-regulation in glioblastoma. In this study, we sought to demonstrate the significance of SLC22A18 as a tumor suppressor in colorectal cancer (CRC) and provide mechanistic bases for its function. We first showed that the expression of SLC22A18 is significantly down-regulated in tumor tissues using matched normal-tumor samples from CRC patients. This finding was also supported by publically accessible data from The Cancer Genome Atlas (TCGA). Functionally, SLC22A18 inhibits colony formation and induces of G2/M arrest consistent with being a tumor suppressor. Interestingly, suppression of KRAS by RNA interference promotes SLC22A18 expression, and expression of SLC22A18 in turn inhibits KRASG12D-mediated anchorage independent growth of NIH3T3 cells indicating a mutual negative interaction. Finally, we evaluated diagnostic and prognostic values of SLC22A18 using clinical and gene expression data from TCGA which revealed a significantly worse long-term prognosis for patients with low level SLC22A18 expression. In sum, we established SLC22A18 as a tumor suppressor in colon epithelial cells and propose that SLC22A18 is potentially a marker of diagnostic and prognostic values.

Silencing of peroxiredoxin II by promoter methylation is necessary for the survival and migration of gastric cancer cells

Peroxiredoxin (Prx), a family of ubiquitous thiol peroxidases, functions as a redox signaling regulator that controls cellular H2O2 in mammalian cells and has recently received attention for being overexpressed in various cancer types. In this study, we show that Prx type II (PrxII) is rather silenced in gastric cancer cells. PrxII expression is severely downregulated in 9 out of the 28 gastric cancer cell lines. Strikingly, PrxII expression is completely lost in three cell lines, MKN28, MKN74 and SNU484. Loss of PrxII expression is due to DNA methyltransferase 1-dependent methylation at the promoter region of the PrxII gene. Restoration of PrxII expression using a retroviral system markedly reduces the colony-forming ability and migratory activity of both MKN28 and SNU484 cells by inhibiting Src kinase. Mechanistically, PrxII peroxidase activity is essential for regulating gastric cancer cell migration. Bioinformatics analysis from The Cancer Genome Atlas stomach cancer data (STAD) revealed significantly low PrxII expression in gastric cancer patients and a negative correlation between PrxII expression and methylation levels. More importantly, low PrxII expression also strongly correlates with poor survival in cancer patients. Thus our study suggests that PrxII may be the first thiol peroxidase that simultaneously regulates both survival and metastasis in gastric cancer cells with high clinical relevance.

GEMiCCL: mining genotype and expression data of cancer cell lines with elaborate visualization

Abstract Cancer cell lines are essential components for biomedical research. However, proper choice of cell lines for experimental purposes is often difficult because genotype and/or expression data are missing or scattered in diverse resources. Here, we report Gene Expression and Mutations in Cancer Cell Lines (GEMiCCL), an online database of human cancer cell lines that provides genotype and expression information. We have collected mutation, gene expression and copy number variation (CNV) data from three representative databases on cell lines—Cancer Cell Line Encyclopedia , Catalogue of Somatic Mutations in Cancer and NCI60. In total, GEMiCCL includes 1406 cell lines from 185 cancer types and 29 tissues. Gene expression, mutation and CNV information are available for 1304, 1334 and 1365 cell lines, respectively. We removed batch effects due to different microarray platforms using the ComBat software and re-processed the entire gene expression and SNP chip data. Cell line names and clinical information were standardized using Cellosaurus from ExPASy. Our user interface supports cell line search, gene search, browsing for specific molecular characteristics and complex queries-based on Boolean logic rules. We also implemented many interactive features and user-friendly visualizations. Providing molecular characteristics and clinical information, we believe that GEMiCCL would be a valuable resource for biomedical research for functional or screening studies. Database URL: GEMiCCL is available at https://www.kobic.kr/GEMICCL/

Characterization of developmental defects in the forebrain resulting from hyperactivated mTOR signaling by integrative analysis of transcriptomic and proteomic data

Hyperactivated mTOR signaling in the developing brain has been implicated in multiple forms of pathology including tuberous sclerosis complex (TSC). To date, various phenotypic defects such as cortical lamination irregularity, subependymal nodule formation, dysmorphic astrocyte differentiation and dendritic malformation have been described for patients and animal models. However, downstream networks affected in the developing brain by hyperactivated mTOR signaling have yet to be characterized. Here, we present an integrated analysis of transcriptomes and proteomes generated from wild-type and Tsc1/Emx1-Cre forebrains. This led to comprehensive lists of genes and proteins whose expression levels were altered by hyperactivated mTOR signaling. Further incorporation of TSC patient data followed by functional enrichment and network analyses pointed to changes in molecular components and cellular processes associated with neuronal differentiation and morphogenesis as the key downstream events underlying developmental and morphological defects in TSC. Our results provide novel and fundamental molecular bases for understanding hyperactivated mTOR signaling-induced brain defects which can in turn facilitate identification of potential diagnostic markers and therapeutic targets for mTOR signaling-related neurological disorders.

Abstract 1532: Exome and transcriptome profiling of lung adenocarcinoma in female never-smokers

Background: Lung cancer is the leading cause of cancer-related mortality in the world. TCGA projects for lung adenocarcinoma and squamous cell carcinoma are the largest datasets publicly available so far. They include diverse subtypes and population groups and thus small population groups such as female never-smokers in Asian are under-represented. Female never-smokers are an important population cohort where mutations due to genetic factors are enriched rather than environmental factors such as smoking. We applied the high throughput deep sequencing strategy to catalog somatic mutations and to profile gene expression levels for Korean patients. Results: Deep sequencing data of exome and transcriptome were generated for tumor and matched normal tissues from 100 non-small-cell lung cancer (NSCLC) patients in female never-smokers. The mutational catalogue includes numerous somatic mutations, indels, and gene-fusion events. Comparison with TCGA lung cancer data shows that our patients harbor much less and homogeneous somatic mutations as expected. Mutational frequency and driver gene analyses identified many novel candidates of causal genetic variations. For example, our mutation spectrum reveals 5 key modules of enriched somatic mutations such as EGFR, p53, Wnt signaling pathways, ECM organization, and cell cycle checkpoints. Furthermore, network analysis of integrating mutation and expression data illustrates the detailed mechanism of dysregulation in tumorigenesis of lung adenocarcinoma. Conclusion: Our study elucidates clinically important mutations and regulatory elements in NSCLC. It is expected that our dataset would serve as an important resource for lung cancer research especially in Asian populations. Citation Format: Sanghyuk Lee, Pora Kim, Yukyung Jun, Charny Park, Suyeon Kim, Hee-Young Lee, Juhee Keum, Yeonhwa Jung, Yeonjoo Jung, Chaehwa Seo, Kyoohyoung Rho, Jong-Eun Lee, Wankyu Kim, Jaesang Kim, Jhingook Kim. Exome and transcriptome profiling of lung adenocarcinoma in female never-smokers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1532. doi:10.1158/1538-7445.AM2014-1532

Colistin therapy for multidrug-resistant Gram-negative infection: clinical outcome and risk factors

Multidrug-resistant (MDR) Gram-negative bacterial infection is widespread all over the world, and recently it led to the reintroduction of colistin for treatment of MDR Pseudomonas aeruginosa and Acinetobacter spp. infection [1]. According to the 2007 European antimicrobial resistance surveillance system (EARSS) consisting of 33 European countries, six countries reported carbapenem resistance rates higher than 25 % among P. aeruginosa isolates [2]. The rate of imipenem-resistant Acinetobacter reached up to 51 % in Korea according to the report of the Korean Nationwide Surveillance of Antimicrobial Resistance (KONSAR) [3]. Polymyxin E (colistin) that was developed in the 1940s is still the principal therapeutic agent for MDR Gram-negative pathogens. Because of the nephrotoxicity and neurotoxicity of colistin, it was not used for a long time [4]. Colistin has lately been considered for MDR treatment due to the emergence of MDR bacteria. However, the efficacy test results of colistin have been quite controversial. Most studies have been retrospective, and the clinical data analysis has been done without control groups [5, 6]. In this study, we evaluated the clinical efficacy of colistin in MDR bacterial infection. We conducted a retrospective hospital-based cohort study at an 850-bed hospital by reviewing the medical records of patients who received intravenous colistin for MDR bacterial infection from May 2007 to December 2010. Demographics, clinical presentation, causative organism, and outcomes were recorded. Patients were excluded if they received colistin treatment for \72 h and were under the age of 18. Types of infection were evaluated, following the United States Centers for Disease Control and Prevention (CDC) criteria. Recent invasive procedure was defined as preceding an invasive procedure within 48 h, including arterial, central venous catheter, urinary catheter, or applying mechanical ventilation before the culture results of MDR infection. MDR bacteria were defined to be resistant to all antibiotic agents such as penicillins, aminoglycosides, ampicillin/sulbactam, cephalosporins, aztreonam, carbapenems, fluoroquinolones, and tetracyclines, except colistin. Interpretation of susceptibility results was carried out according to the current Clinical and Laboratory Standards Institute guidelines. A total of 107 patients were enrolled in this study, and their demographic and clinical data are summarized in Table 1. Sixty-six patients (61.7 %) were over 64 years, and 36 patients (33.6 %) had a hospital stay of more than 125 days before MDR bacteria culture. Common underlying diseases were hypertension (63.5 %), diabetes (34.6 %), chronic obstructive pulmonary disease (24.3 %), cancer (19.6 %), chronic renal failure (CRF), and end-stage renal disease (ESRD; 12.1 % for both CRF and ESRD). More than 95 % of colistin therapy took place in ICUs. Cerebrovascular event (27.1 %) including cerebral infarction or hemorrhage was the most common cause of admission. We evaluated the clinical and microbiological responses both to colistin and to previously exposed antibiotics combined with colistin. Efficacy was evaluated as both clinical and bacteriological responses. After 3 days of colistin therapy, clinical improvement was graded as a good or poor clinical response. A good responder was defined as a patient whose symptoms were improved with bacteriologically clear pathogen within 1 week. Baseline characteristics and various risk factors were compared Y. H. Jun S. J. Jeun S. H. Kang H. J. Choi (&) Division of Infectious Disease, Department of Internal Medicine, Ewha Womans University, Mokdong Hospital, 911-1 MokDong, Yangcheon-Ku, Seoul, Korea e-mail: heechoi@ewha.ac.kr