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Gut microbiota provide numerous types of metabolites that humans cannot produce and have a huge influence on the host metabolism. Accordingly, gut bacteria-derived metabolites can be employed as a resource to develop anti-obesity and metabolism-modulating drugs.This study aimed to examine the anti-adipogenic effect of 3-phenylpropionylglycine (PPG), which is a glycine conjugate of bacteria-derived 3-phenylpropionic acid (PPA).The effect of PPG on preadipocyte-to-adipocyte differentiation was evaluated in 3T3-L1 differentiation models and the degree of the differentiation was estimated by Oil red O staining. The molecular mechanisms of the PPG effect were investigated with transcriptome analyses using RNA-sequencing and quantitative real-time PCR.PPG suppressed lipid droplet accumulation during the adipogenic differentiation of 3T3-L1 cells, which is attributed to down-regulation of lipogenic genes such as acetyl CoA carboxylase 1 (Acc1) and fatty acid synthase (Fasn). However, other chemicals with chemical structures similar to PPG, including cinnamoylglycine and hippuric acid, had little effect on the lipid accumulation of 3T3-L1 cells. In transcriptomic analysis, PPG suppressed the expression of adipogenesis and metabolism-related gene sets, which is highly associated with downregulation of the peroxisome proliferator-activated receptor (PPAR) signaling pathway. Protein-protein association network analysis suggested adiponectin as a hub gene in the network of genes that were differentially expressed genes in response to PPG treatment.PPG inhibits preadipocyte-to-adipocyte differentiation by suppressing the adiponectin-PPAR pathway. These data provide a potential candidate from bacteria-derived metabolites with anti-adipogenic effects.

Metastatic or recurrent colorectal cancer (CRC) patients require systemic chemotherapy, but the therapeutic options of targeted agents remain limited. CRC patients with KRAS or BRAF gene mutations exhibit a worse prognosis and are resistant to anti-EGFR treatment. Previous studies have shown that the expression of anti-apoptotic protein BCL-XL is increased in CRC patients with KRAS/BRAF mutations, suggesting BCL-XL as a therapeutic target for this subgroup. Here, we performed genome-wide CRISPR/Cas9 screens of cell lines with KRAS mutations to investigate the factors required for sensitivity to BCL-XL inhibitor ABT-263 using single-guide RNAs (sgRNAs) that induce loss-of-function mutations. In the presence of ABT-263, sgRNAs targeting negative regulators of WNT signaling (resulting in WNT activation) were enriched, whereas sgRNAs targeting positive regulators of WNT signaling (resulting in WNT inhibition) were depleted in ABT-263-resistant cells. The activation of WNT signaling was highly associated with an increased expression ratio of anti- to pro-apoptotic BCL-2 family genes in CRC samples. Genetic and pharmacologic inhibition of WNT signaling using β-catenin short hairpin RNA or TNIK inhibitor NCB-0846, respectively, augmented ABT-263-induced cell death in KRAS/BRAF-mutated cells. Inhibition of WNT signaling resulted in transcriptional repression of the anti-apoptotic BCL-2 family member, MCL1, via the functional inhibition of the β-catenin-containing complex at the MCL1 promoter. In addition, the combination of ABT-263 and NCB-0846 exhibited synergistic effects in in vivo patient-derived xenograft (PDX) models with KRAS mutations. Our data provide a novel targeted combination treatment strategy for the CRC patient subgroup with KRAS or BRAF mutations.

N6-methyladenosine RNA methylation is one of the most abundant post transcriptional modifications in eukaryotic messenger RNAs and plays crucial roles in cancer development and progression. YTHDF1 is an m6A reader protein and has been reported to increase the translation of mRNAs in cooperation with YTHDF3. In gastric cancer, YTHDF1 was reported to be overexpressed in cancer tissues, but detailed effects of YTHDF1 in cancer immunity have not been fully elucidated. In RNA sequencing data from a TGCA gastric cancer cohort, the expression of YTHDF1 was inversely correlated with the cytotoxic T cell markers, such as CD8A, GZMA, and GZMB. To investigate the effect of YTHDF1 overexpression on gene expression regulation in protein levels, we performed the proteomic analysis for newly synthesized proteins using non-canonical amino acid azidohomoalanine and click chemistry tagging reaction in the presence of interferon-γ (IFN-γ). Protein synthesis of genes in ‘DNA damage response’ and ‘DNA repair’ gene sets increased in YTHDF1-overexpressed samples. On the contrary, protein synthesis of genes in ‘Allograph rejection’, ‘NK cell mediated cytotoxicity’, and ‘NF-κB signaling’ gene sets decreased in YTHDF1-overexpressed samples. Especially, YTHDF1 reduced basal expression of DNA damage marker, γH2AX, suggesting that DNA damage repair was enhanced by YTHDF1 overexpression. Our study suggests that YTHDF1 plays pervasive roles in DNA repair and cancer immunity by regulating gene expression in protein levels. Citation Format: Dongjun Jang, Hae Rim Jung, Jaeik Oh, Sung-Yup Cho. RNA N6-methyladenosine binding protein, YTHDF1 regulates DNA repair and cancer immunity in protein synthesis levels. [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 3716.

Purpose The probability of recurrence of cancer after adjuvant or definitive radiotherapy in patients with human papillomavirus-negative (HPV(-)) head and neck squamous cell carcinoma (HNSCC) varies for each patient. This study aimed to identify and validate radiation sensitivity signature (RSS) of patients with HPV(-) HNSCC to predict the recurrence of cancer after radiotherapy. Materials and methods Clonogenic survival assays were performed to assess radiosensitivity in 14 HNSCC cell lines. We identified genes closely correlated with radiosensitivity and validated them in The Cancer Genome Atlas (TCGA) cohort. The validated RSS were analyzed by ingenuity pathway analysis (IPA) to identify canonical pathways, upstream regulators, diseases and functions, and gene networks related to radiosensitive genes in HPV(-) HNSCC. Results The survival fraction of 14 HNSCC cell lines after exposure to 2 Gy of radiation ranged from 48% to 72%. Six genes were positively correlated and 35 genes were negatively correlated with radioresistance, respectively. RSS was validated in the HPV(-) TCGA HNSCC cohort (n = 203), and recurrence-free survival (RFS) rate was found to be significantly lower in the radioresistant group than in the radiosensitive group (p = 0.035). Cell death and survival, cell-to-cell signaling, and cellular movement were significantly enriched in RSS, and RSSs were highly correlated with each other. Conclusion We derived a HPV(-) HNSCC-specific RSS and validated it in an independent cohort. The outcome of adjuvant or definitive radiotherapy in HPV(-) patients with HNSCC can be predicted by analyzing their RSS, which might help in establishing a personalized therapeutic plan.

N6-Methyladenosine (m6A) RNA modification plays a critical role in the posttranscriptional regulation of gene expression. Alterations in cellular m6A levels and m6A-related genes have been reported in many cancers, but whether they play oncogenic or tumor-suppressive roles is inconsistent across cancer types. We investigated common features of alterations in m6A modification and m6A-related genes during carcinogenesis by analyzing transcriptome data of 11 solid tumors from The Cancer Genome Atlas database and our in-house gastric cancer cohort. We calculated m6A writer (W), eraser (E), and reader (R) signatures based on corresponding gene expression. Alterations in the W and E signatures varied according to the cancer type, with a strong positive correlation between the W and E signatures in all types. When the patients were divided according to m6A levels estimated by the ratio of the W and E signatures, the prognostic effect of m6A was inconsistent according to the cancer type. The R and especially the R2 signatures (based on the expression of IGF2BPs) were upregulated in all cancers. Patients with a high R2 signature exhibited poor prognosis across types, which was attributed to enrichment of cell cycle- and epithelial–mesenchymal transition-related pathways. Our study demonstrates common features of m6A alterations across cancer types and suggests that targeting m6A R proteins is a promising strategy for cancer treatment.

Metastatic or recurrent colorectal cancer (CRC) patients require systemic chemotherapy, but the therapeutic options of targeted agents remain limited. CRC patients with KRAS or BRAF gene mutations exhibit a worse prognosis and are resistant to anti-EGFR treatment. Previous studies have shown that the expression of anti-apoptotic protein BCL-X

Cancer chemotherapeutic drugs exert cytotoxic effects by modulating intracellular reactive oxygen species (ROS) levels. However, whether ROS modulates the efficacy of targeted therapeutics remains poorly understood. Previously, we reported that upregulation of the anti-apoptotic protein, BCL-X

Objectives: This study investigated whether the biomarkers present in nasal fluid reflect the severity of symptoms in patients with persistent allergic rhinitis (PAR). Methods: We enrolled 29 PAR patients complaining of nasal symptoms and testing positive to skin prick test. Patients’ total nasal symptom score (TNSS) was measured and their nasal lavage fluid (NALF) was collected. The levels of biomarkers including Clara cell protein 16 (CC16), tryptase, and interleukin 5 (IL-5) in NALF were determined via enzyme-linked immunosorbent assay (ELISA). Results: PAR patients were classified into persistent mild and persistent moderate-to-severe groups according to the Allergic Rhinitis and its Impact on Asthma (ARIA) guidelines. The CC16 alone was significantly negatively correlated with TNSS ( P Conclusions: The levels of CC16 alone among several NALF biomarkers showed an inverse correlation with symptoms of PAR patients.

이 연구는 고지혈증에 대한 생리 반응의 조절과 관련하여 Dendropanax morbifera 잎의 효과를 비교하기 위한 것이다. 추출물은 표본 연령, 수확시기, 추출 방법에 따라 달랐다. 이 추출물이 처리된 RAW 264.7 세포는 산화 저밀도 지단백질에 의해 자극되었고, 세포 지질 축적은 Oil Red O (ORO) 염색법을 사용하여 정량화되었다. HepG2 세포에서 에탄올을 사용하여 독성을 유도한 다음, 상기와 같이 지질 축적을 정량화하였다. 추출물의 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase 억제 활성을 측정하였다. RAW 264.7 세포에서 가장 지질 축적을 억제한 시료는 8년생 11월 채취 주정 60% 추출물로 처리된 세포에서 관찰되었다. 에탄올로 유도한 HepG2 세포에서 가장 큰 보호효과를 나타낸 시료는 5년생 5월 채취 주정30% 추출물이었다. 가장 강력한 HMG-CoA reductase 억제 작용을 한 시료는 8년생 5월 채취 열수 100% 추출물이다. 전체적으로 봤을 때 지질 축적 억제는 주정 추출물에서, 콜레스테롤 합성 효소 저해 효과는 열수추출물에서 좋은 효과를 보여 이들 원료를 기반으로 지질 대사 개선 기능성 원료를 개발할 수 있을 것이라 기대한다.

This study compared the antioxidant activity of extracts from Dendropanax morbifera (D. morbifera) Levillis leaves. The concentrations of flavonoids and polyphenols were measured in extracts of D. morbifera leaves. The antioxidant activities were examined by ABTS and DPPH radical scavenging activity and ferric reducing antioxidant power (FRAP). Total flavonoid and polyphenol contents, and FRAP were highest in the 30% ethanol extract collected in May. The ABTS and DPPH radical scavenging activities were the highest in the 60% ethanol extract harvested in May. For investigating the relationship between antioxidant activity and specific polyphenols, rutin and chlorogenic acid of the polyphenol component were quantified by LC–MS/MS analysis. The concentrations of them were highest in the 60% ethanol extract collected in May, and showed positive correlations with antioxidant activities. The optimal extraction conditions to yield the most effective antioxidant activity were obtained using a 60% ethanol extraction solvent with samples collected in May.

SOX2 copy number and mRNA expression were analysed to examine the clinical significance of SOX2 activation in HNSCC. Gene expression signatures reflecting SOX2 activation were identified in an HNSCC cohort. Patients with HNSCC were classified into two subgroups according to the gene expression signature: SOX2-high and SOX2-low. The clinical significance of SOX2 activation was further validated in two independent cohorts. Moreover, clinical significance of SOX2 activation in response to radiotherapy was assessed in patients with HNSCC. The relationship between SOX2 activation and radiotherapy was validated in an in vitro experiment. Patients in the SOX2-high subgroup had a better prognosis than patients in the SOX2-low subgroup in all three patient cohorts. Results of multivariate regression analysis showed that SOX2 signature was an independent predictor of the overall survival of patients with HNSCC (hazard ratio, 1.45; 95% confidence interval, 1.09–1.92; P = 0.01). Interestingly, SOX2 activation was a predictor of therapy outcomes in patients receiving radiotherapy. Moreover, SOX2 overexpression enhanced the effect of radiotherapy in HNSCC cell lines. SOX2 activation is associated with improved prognosis of patients with HNSCC and might be used to predict which patients might benefit from radiotherapy.

// Hae Ryung Chang 1, 2, * , Seungyoon Nam 1, 3, * , Jinhyuk Lee 4, 5, * , Jin-Hee Kim 6, * , Hae Rim Jung 1 , Hee Seo Park 7 , Sungjin Park 1, 3 , Young Zoo Ahn 1 , Iksoo Huh 8 , Curt Balch 9 , Ja-Lok Ku 10 , Garth Powis 11 , Taesung Park 8 , Jin-Hyun Jeong 6 , Yon Hui Kim 1, 12 1 New Experimental Therapeutics Branch, National Cancer Center of Korea, Goyang-si, Republic of Korea 2 Research Institute of Women’s Health, Sookmyung Women’s University, Seoul, Republic of Korea 3 College of Medicine, Gachon University, Incheon, Republic of Korea 4 Korean Bioinformation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea 5 Department of Nanobiotechnology and Bioinformatics, Korea University of Science and Technology, Daejeon, Republic of Korea 6 College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, Incheon, Republic of Korea 7 Animal Sciences Branch, National Cancer Center of Korea, Goyang-si, Republic of Korea 8 Department of Statistics, Seoul National University, Seoul, Republic of Korea 9 Bioscience Advising, Ypsilanti, MI, USA 10 Korean Cell Line Bank, Seoul National University, Seoul, Republic of Korea 11 Cancer Center, Sanford-Burnham Prebys Medical Discovery Institute, La Jolla, CA, USA 12 Discovery Biology, CrystalGenomics Inc., Seongnam-si, Republic of Korea * These authors have contributed equally to this work Correspondence to: Yon Hui Kim, email: yhkim@cgxinc.com Jin-Hyun Jeong, email: organicjeong@yonsei.ac.kr Taesung Park, email: tspark@stats.snu.ac.kr Keywords: gastric cancer, RHOA, G-protein, biomarker, therapeutic target Received: July 11, 2016 Accepted: October 17, 2016 Published: October 28, 2016 ABSTRACT Gastric cancer (GC) is a highly heterogeneous disease, in dire need of specific, biomarker-driven cancer therapies. While the accumulation of cancer “Big Data” has propelled the search for novel molecular targets for GC, its specific subpathway and cellular functions vary from patient to patient. In particular, mutations in the small GTPase gene RHOA have been identified in recent genome-wide sequencing of GC tumors. Moreover, protein overexpression of RHOA was reported in Chinese populations, while RHOA mutations were found in Caucasian GC tumors. To develop evidence-based precision medicine for heterogeneous cancers, we established a systematic approach to integrate transcriptomic and genomic data. Predicted signaling subpathways were then laboratory-validated both in vitro and in vivo , resulting in the identification of new candidate therapeutic targets. Here, we show: i) differences in RHOA expression patterns, and its pathway activity, between Asian and Caucasian GC tumors; ii) in vitro and in vivo perturbed RHOA expression inhibits GC cell growth in high RHOA-expressing cell lines; iii) inverse correlation between RHOA and RHOB expression; and iv) an innovative small molecule design strategy for RHOA inhibitors. In summary, RHOA, and its oncogenic signaling pathway, represent a strong biomarker-driven therapeutic target for Asian GC. This comprehensive strategy represents a promising approach for the development of “hit” compounds.

The proximate composition and various specific components of Antarctic krill Euphausia superba , in the catch season between March and August were investigated. Frozen krill were freeze-dried and milled. The proximate composition comprised water, proteins, fats, ash, fatty acids, and amino acids, while the specific components were vitamins, minerals, nucleotides, betaine, and astaxanthin. The moisture content of the krill ranged from 77 to 80%, with the highest value in June, and the ash content was be-tween 12 and 13%. The protein content was lowest in May, and the fat content was 18–19%, with the highest value in March. The amino acid content varied according to the season: taurine and glycine were highest in August; β-alanine was higher in April and May; and arginine, ornithine, and lysine were highest in March. The unsaturated fat content was ~50% and omega-3 fatty acids were highest in June. Oil-soluble vitamins A and E were highest in March, and the water-soluble vitamin content was less than that of oil-soluble vitamins. The mineral content was highest in June, and the most abundant mineral was sodium at 235.60 mg/100 g krill. The content of other minerals was lowest (2.94 mg/100 g) in April, except for lead. The nucleotide content was highest in July, while the betaine content was highest in April and lowest in June. The astaxanthin content was highest in May and ranged from 6 to 10 ppm in other months.

Background Worldwide, gastric cancer (GC) is the fourth most common malignancy and the most common cancer in East Asia. Development of targeted therapies for this disease has focused on a few known oncogenes but has had limited effects. Objective To determine oncogenic mechanisms and novel therapeutic targets specific for GC by identifying commonly dysregulated genes from the tumours of both Asian-Pacific and Caucasian patients. Methods We generated transcriptomic profiles of 22 Caucasian GC tumours and their matched non-cancerous samples and performed an integrative analysis across different GC gene expression datasets. We examined the inhibition of commonly overexpressed oncogenes and their constituent signalling pathways by RNAi and/or pharmacological inhibition. Results Hepatocyte nuclear factor-4α (HNF4α) upregulation was a key signalling event in gastric tumours from both Caucasian and Asian patients, and HNF4α antagonism was antineoplastic. Perturbation experiments in GC tumour cell lines and xenograft models further demonstrated that HNF4α is downregulated by AMPKα signalling and the AMPK agonist metformin; blockade of HNF4α activity resulted in cyclin downregulation, cell cycle arrest and tumour growth inhibition. HNF4α also regulated WNT signalling through its target gene WNT5A, a potential prognostic marker of diffuse type gastric tumours. Conclusions Our results indicate that HNF4α is a targetable oncoprotein in GC, is regulated by AMPK signalling through AMPKα and resides upstream of WNT signalling. HNF4α may regulate ‘metabolic switch’ characteristic of a general malignant phenotype and its target WNT5A has potential prognostic values. The AMPKα-HNF4α-WNT5A signalling cascade represents a potentially targetable pathway for drug development.

The translation of high-throughput gene expression data into biologically meaningful information remains a bottleneck. We developed a novel computational algorithm, PATHOME, for detecting differentially expressed biological pathways. This algorithm employs straightforward statistical tests to evaluate the significance of differential expression patterns along subpathways. Applying it to gene expression data sets of gastric cancer (GC), we compared its performance with those of other leading programs. Based on a literature-driven reference set, PATHOME showed greater consistency in identifying known cancer-related pathways. For the WNT pathway uniquely identified by PATHOME, we validated its involvement in gastric carcinogenesis through experimental perturbation of both cell lines and animal models. We identified HNF4α-WNT5A regulation in the cross-talk between the AMPK metabolic pathway and the WNT signaling pathway, and further identified WNT5A as a potential therapeutic target for GC. We have demonstrated PATHOME to be a powerful tool, with improved sensitivity for identifying disease-related dysregulated pathways.

Instant krill gruel was produced using peeled Antarctic krill Euphausia superba as a high nutritional ingredient and the food quality was investigated. The food quality of krill gruel was examined by measuring proximate composition, cholesterol, calorie, Hunter color value, total amino acids, fatty acids, fluoride, viscoelastic properties, and by sensory evaluation. The krill gruel had a moisture content of 87% and a pH of 6.65. The krill gruel contained 51 kcal/100 g, and 0.1% fat and 3.5 mg/100 g cholesterol. Its fatty acid composition exhibited high levels of unsaturated fatty acids. The levels of oleic acid and linolenic acid were high, and n-3, n-6, and n-9 fatty acid contents ranged from 1% to 6%. The total amino acid content was 2132 mg/100 g, and the levels of glutamic acid, aspartic acid, leucine, alanine, and arginine were particularly high. Essential amino acids accounted for over 30% of the total amino acids. Fluoride level in the krill gruel was 3.07 mg/kg. The viscoelastic properties of the krill gruel were determined as 6.28 Pa at shear stress of 2.51 Pa. In the recovery test, the elastic restoring force after deformation was low.

In the process of developing individualized cancer therapeutics, tumor derived cell line based preclinical (in vitro and in vivo) methods have been widely used. Patient derived genome analysis revealed significant cancer biomarkers for targeted drug development. Classic cancer biomarker, HER2, overexpression was discovered as a cancer biomarker and as a druggable target in breast cancer, which lead to the development of Trastuzumab. HER2 is overexpressed in 10-25% of gastric cancer, and treatment with Trastuzumab showed positive yet limited activity in advanced gastric cancer treatment in the ToGA study. These results present HER2 as an attractive target for gastric cancer. However, Gillet and colleagues reported the challenges in the use of in vitro and in vivo studies for drug validation and development. They reported variance in gene expression patterns between the in vitro and in vivo context within a single cancer cell line. In our study using gastric cancer derived cell lines, we observed differences in HER2 expression between in vitro and in vivo xenograft model. Also, we show the potential promising in vivo xenograft model in the development of HER2 targeting agents for gastric cancer. Eleven out of 25 gastric cancer cell lines showed HER2 expression, which was demonstrated using the IHC assay on a cell microarray. Sixteen out of 25 cell lines successfully produced xenograft models. Gastric cancer cell lines (SNU-5, SNU-16, SNU-638, SNU-1967, MKN-1, MKN-28, and NUGC-3) that had no HER2 expression when cultured in vitro showed HER2 expression in vivo. Two cell lines (NCC-19 and NCI-N87) that had HER2 expression in vitro showed a decrease in HER2 expression in vivo. Three cell lines (SNU-484, SNU-668, and SNU-719) showed various levels of increase in HER2 expression. Two cell lines showed minor to no changes in HER2 expression. NUGC-4 had consistent significant HER2 expression levels in both assays, while MKN-45 had consistent insignificant to no HER2 expression. Our study demonstrates inconsistency in HER2 gene expression between in vitro and in vivo cancer cell line models for the first time in gastric cancer. Citation Format: Youme Gim, Hee Seo Park, Hae Rim Jung, Hae Ryung Chang, Seungyoon Nam, Yong Doo Choi, Ja-Lok Ku, Yon Hui Kim. Variability of HER2 expression between in vitro and in vivo models within gastric cancer cell lines. [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 2783. doi:10.1158/1538-7445.AM2014-2783

Gastric cancer(GC) is the 4th most common malignancy in the world and the 2nd most common cause of cancer related death worldwide (1). The known causes of GC are diet, hygiene, Helicobacter pylori, Epstein-Barr virus and hereditary factors. Other genetic causes of GC are known to be heterogenic, partially due to differences in ethnic/genetic background. Surgical removal of the tumor is the first-line treatment. Targeted therapy agents and chemotherapeutic agents that show survival advantage in other cancer types are being evaluated in GC(2,3), which only target few known oncogenes. Targeted therapy for GC is still an unmet need, and in light of personalized medicine, identifying genes specific to GC is critical for its treatment. From our NGS data, we have discovered protein family members involved in focal adhesion and actin cytoskeleton regulation pathways, which are involved in mitosis and metastasis. We developed a subpathway network analysis method called PATHOME(unpublished), to identify pathways related to GC. For the first stage, two datasets (84 Korean GC: GSE13861 and 56 Japanese GC:GSE15081)(4,5). Two datasets (25 Korean GC: GSE36968 and 160 Chinese GC:GSE27342)(6,7,8) were used as validation sets. We identified top 10 common pathways. We have then categorized genes within these pathways according their functional category and druggability, either as drug transporter or druggable target. As a result, we identified genes involved in the focal adhesion and regulation of actin cytoskeleton pathways and cell junction, all critical for cell division and metastasis of tumor cells. Genes G1-9 are tight junction genes, and G10-20 are protein family group related to Ras-like GTP-binding protein, G10 - 20. G10, 11 and 12 are ras-like GTPases, and G13-20 are interacting proteins or downstream proteins. We then compared differential expression of these genes between the Asian and Caucasian dataset. G1-9 show co-regulation pattern among the two ethnic group. G9 in particular, is normally expressed in the stomach, and is downregulated (p=0.0045) in GC tumor samples. In contrast, G10 is upregulated in the Korean dataset and is shown to be upregulated in Japanese and Chinese GC sample (public) as well(9,10), but not in our Caucasian dataset(unpublished). The TCGA stomach cancer provisional data via cBioPortal(11) shows that most of the G10 alterations in GC cases are mutations (9% of total cases) with 2 known cases of downregulation. It is interesting that most of the G10 alterations were found in Caucasian patients (11/18 cases). In our study, we have identified genes that are significant in GC and are druggable targets/drug transporters. It is significant that the pathways we identified are detected in both ethnic groups, with different expression pattern. This finding is critical for personalized treatment and drug development for GC. Citation Format: Hae Ryung Chang, Seungyoon Nam, Myeong-Cherl Kook, Hee Seo Park, Hae Rim Jung, Youme Gim, Han Liang, Garth Powis, Yon Hui Kim. Identification of focal adhesion and actin cytoskeleton regulation family genes as druggable target for gastric cancer. [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 1781. doi:10.1158/1538-7445.AM2014-1781

The LKB1/AMPK signaling pathway has been well elucidated and recent evidence suggests its involvement in cancer cell biology, demonstrating that the reinforcement of the tumor suppressive functions of LKB1/AMPK is a valuable therapeutic strategy for cancers. Interest in metformin as a novel anticancer agent for breast cancer and other solid tumors continue to grow, currently being investigated in several cancer types in both neoadjuvant and metastatic settings. The biological effect of metformin on cancer cells is driven by its ability to activate AMPK through upstream kinase LKB1, tumor suppressor gene in epithelial tissues. Metformin increases intracellular AMP level, which allosterically activates AMPK. We have previously identified the AMPKα as a modulator in gastric cancer (GC) and through experimental evidence. We show the impact of LKB1/AMPK modulation of HNF4α, a dramatic suppression of cancer cell growth. GC samples were collected and sequenced on SOLiD v 3.0 for both WT-seq and small RNA-seq. Computational analysis showed that 356 out of 18,890 genes were identified as GC related differentially expressed genes in the five-group comparison (normal, tumor stage I, II, III or IV). 28 genes were identified as stage-specific differentially expressed genes, and 13 out of the 28 genes were within the network between HNF4α and HNF1α. In order to test the anti-proliferation activity of metformin in GC cell lines associated with activation of PRKAA1, PRKAA2 and LKB1, and by HNF4α suppression, 4 GC cell lines (NCI-N87, AGS, HS 746T and MKN 45) were treated with metformin. Both PRKAA1/2 showed increased gene expression level when the cells were treated with 10mM of metformin. As for STK11 (LKB1 gene) and HNF4A gene expression level, LKB1 increased and HNF4A decreased with metformin treatment on all four cell lines. Metformin treated NCI-N87 and AGS show that it is involved in cell cycle arrest. Western-blot analysis shows, decreased protein expression of Cyclin A/B and D1 on metformin treated. Lastly, NCI-N87 xenograft study show metformin treated suppression of tumor progression compared to non-treated mouse. During the 28 day treatment of metformin, PRKAA1 and PRKAA2 expression level increased compared to the untreated with. Consistent with in vitro assay, LKB1 level was elevated in the metformin treated tumor compared to the non-treated, and HNF4α level decreased over time in the metformin treated tumor. Study shows that AMPK is a strong therapeutic tumor suppressor target and that metformin is a potential drug for Asian early gastric cancer patient. In our research in progress, we observe potential relationships between the Wnt pathway and AMPKα in light of WNT druggability with metformin. In conclusion LKB1/AMPK by HNF4α inhibition suggests metformin could be a candidate for gastric cancer treatment, probably in combination with conventional chemotherapy and/or as a maintenance therapy. Citation Format: Hae Rim Jung, Hae Ryung Chang, Hye-Hyun Seo, Robert Lemos, Hee Seo Park, Han Liang, Garth Powis, Yon Hui Kim. Metformin increases AMPKα activity by inhibition of AMPKα and cell cycle proliferation in Asian gastric cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5534. doi:10.1158/1538-7445.AM2013-5534

Stomach cancer is the most prevalent cancer in Eastern Asia(1). Given the diverse route to oncognesis, it is highly attractive to develop targeted therapy based on each patient's biology(2). Several molecular targeting agents that show survival advantage in other cancer types are being under clinical investigation in gastric cancer (GC) (3-6). These agents focus on a few known oncogenic genes, yet its true efficacy of their biologically targeted therapies in gastric cancer is still unknown. To identify high-quality therapeutic targets, we utilized PATHOME (pathway and genome information), a computational method to identify therapeutic target through sub-pathway employing simple statistic-based approach (unpublished). PATHOME design had two stages: 1) discovery stage aimed at selecting target signaling pathway, and 2) validation stage aimed at identifying small number of reproducible genes from sub-pathways by using independent dataset to provide a novel theragnostic strategy in GC patients. Four sets of GEO database datasets were used: GSE13861 (84 Korean GC), GSE15081 (56 Japanese GC) for the discovery stage, and GSE36968 (25 Korean GC), and GSE27342 (160 Chinese GC) were added for validation. Using Vogelstein et al. as the gold standard, DAVID and GSEA vs. PATHOME were applied for comparison of potency in prediction of signaling pathways(7). Previous study showed that the energy sensing mechanism by AMPKα is involved in Asian GC(8). Metformin is a drug used to treat diabetes, which affects AMPKα in the cell. We treated GC cell line with metformin and saw the expression level of proteins that were selected by PATHOME to test its predictability of biological context. 13 up-regulated genes were identified using 3 datasets (GSE 13861, GSE 15081 and GSE36968), and 5 (GENE1, GENE2, GENE3, GENE4 and GENE5) from all 4 datasets using PATHOME. Transcription factor binding site analysis showed that two of the 5 genes contained GENE6 binding sites. Implications of AMPKα as a potential therapeutic target has been published(9). Metformin is shown to increase PRKAA2 level, which encodes AMPKα, also inhibits HNF4α(8). To see if metformin has any effect on the genes with HNF4α binding site, we observed the expression level of GENE4 and related proteins. Metformin treated GC cell lines (NCI-N87 and AGS) result in decreased viability compared to untreated cells. On Western-blot analysis, metformin treated cells show a decrease of GENE4, GENE6, but not GENE7 or GENE8. Utilizing PATHOME, which showed greater consistency compared to DAVID and GSEA in terms of cancer-related pathways, we showed Wnt pathway to have a potential impact on GC in East Asia. Therefore, our approach has simplified insight of multiplexed signals apply in molecular targeted therapies. We report a cross-talk between AMPKα and Wnt pathway in gastric cancer, which is known to be involved in other cancer types. This finding will lead to other potential drug targets for GC. Citation Format: Hae Ryung Chang, Seungyoon Nam, Hae Rim Jung, ChangHyuk Kwon, Hye-Hyun Seo, Frances S. Sung, Hee Seo Park, Taesung Park, Yon Hui Kim. Crosstalk between Wnt and AMPK pathway in Asian gastric cancer by metformin. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5540. doi:10.1158/1538-7445.AM2013-5540

Although trastuzumab is a successful targeted therapy for breast cancer patients with tumors expressing HER2 (ERBB2), many patients eventually progress to drug resistance. Here, we identified subpathways differentially expressed between trastuzumab-resistant vs. -sensitive breast cancer cells, in conjunction with additional transcriptomic preclinical and clinical gene datasets, to rigorously identify overexpressed, resistance-associated genes. From this approach, we identified 32 genes reproducibly upregulated in trastuzumab resistance. 25 genes were upregulated in drug-resistant JIMT-1 cells, which also downregulated HER2 protein by >80% in the presence of trastuzumab. 24 genes were downregulated in trastuzumab-sensitive SKBR3 cells. Trastuzumab sensitivity was restored by siRNA knockdown of these genes in the resistant cells, and overexpression of 5 of the 25 genes was found in at least one of five refractory HER2 + breast cancer. In summary, our rigorous computational approach, followed by experimental validation, significantly implicate ATF4, CHEK2, ENAH, ICOSLG, and RAD51 as potential biomarkers of trastuzumab resistance. These results provide further proof-of-concept of our methodology for successfully identifying potential biomarkers and druggable signal pathways involved in tumor progression to drug resistance.

Background “Biomarker-driven targeted therapy,” the practice of tailoring patients’ treatment to the expression/activity levels of disease-specific genes/proteins, remains challenging. For example, while the anti-ERBB2 monoclonal antibody, trastuzumab, was first developed using well-characterized, diverse in vitro breast cancer models (and is now a standard adjuvant therapy for ERBB2-positive breast cancer patients), trastuzumab approval for ERBB2-positive gastric cancer was largely based on preclinical studies of a single cell line, NCI-N87. Ensuing clinical trials revealed only modest patient efficacy, and many ERBB2-positive gastric cancer (GC) patients failed to respond at all (i.e., were inherently recalcitrant), or succumbed to acquired resistance. Method To assess mechanisms underlying GC insensitivity to ERBB2 therapies, we established a diverse panel of GC cells, differing in ERBB2 expression levels, for comprehensive in vitro and in vivo characterization. For higher throughput assays of ERBB2 DNA and protein levels, we compared the concordance of various laboratory quantification methods, including those of in vitro and in vivo genetic anomalies (FISH and SISH) and xenograft protein expression (Western blot vs. IHC), of both cell and xenograft (tissue-sectioned) microarrays. Results The biomarker assessment methods strongly agreed, as did correlation between RNA and protein expression. However, although ERBB2 genomic anomalies showed good in vitro vs. in vivo correlation, we observed striking differences in protein expression between cultured cells and mouse xenografts (even within the same GC cell type). Via our unique pathway analysis, we delineated a signaling network, in addition to specific pathways/biological processes, emanating from the ERBB2 signaling cascade, as a potential useful target of clinical treatment. Integrated analysis of public data from gastric tumors revealed frequent (10 – 20 %) amplification of the genes NFKBIE, PTK2, and PIK3CA, each of which resides in an ERBB2-derived subpathway network. Conclusion Our comprehensive bioinformatics analyses of highly heterogeneous cancer cells, combined with tumor “omics” profiles, can optimally characterize the expression patterns and activity of specific tumor biomarkers. Subsequent in vitro and in vivo validation, of specific disease biomarkers (using multiple methodologies), can improve prediction of patient stratification according to drug response or nonresponse. Electronic supplementary material The online version of this article (doi:10.1186/s12885-016-2232-2) contains supplementary material, which is available to authorized users.