Tae-Su Han

MicroRNA-29c mediates initiation of gastric carcinogenesis by directly targeting ITGB1

Objective Gastric cancer (GC) remains difficult to cure due to heterogeneity in a clinical challenge and the molecular mechanisms underlying this disease are complex and not completely understood. Accumulating evidence suggests that microRNAs (miRNAs) play an important role in GC, but the role of specific miRNAs involved in this disease remains elusive. We performed next generation sequencing (NGS)-based whole-transcriptome profiling to discover GC-specific miRNAs, followed by functional validation of results. Design NGS-based miRNA profiles were generated in matched pairs of GCs and adjacent normal mucosa (NM). Quantitative RT-PCR validation of miR-29c expression was performed in 274 gastric tissues, which included two cohorts of matched GC and NM specimens. Functional validation of miR-29c and its gene targets was undertaken in cell lines, as well as K19-C2mE and K19-Wnt1/C2mE transgenic mice. Results NGS analysis revealed four GC-specific miRNAs. Among these, miR-29c expression was significantly decreased in GC versus NM tissues (p<0.001). Ectopic expression of miR-29c mimics in GC cell lines resulted in reduced proliferation, adhesion, invasion and migration. High miR-29c expression suppressed xenograft tumour growth in nude mice. Direct interaction between miR-29c and its newly discovered target, ITGB1, was identified in cell lines and transgenic mice. MiR-29c expression demonstrated a stepwise decrease in wild type hyperplasia-dysplasia cascade in transgenic mice models of GC. Conclusions MiR-29c acts as a tumour suppressor in GC by directly targeting ITGB1. Loss of miR-29c expression is an early event in the initiation of gastric carcinogenesis and may serve as a diagnostic and therapeutic biomarker for patients with GC.

Dissemination of Free Cancer Cells from the Gastric Lumen and from Perigastric Lymphovascular Pedicles during Radical Gastric Cancer Surgery

BackgroundManipulation and improper handling of a tumor during surgery may increase the risk of cancer cell dissemination after a curative gastrectomy. This study investigated the effect of improper handling of lymphovascular pedicles of stomach on tumor spillage during surgical procedure.MethodsThirty-eight gastric cancer patients were enrolled. Three pairs of wash samples were obtained from each patient: (1) intraperitoneal wash samples obtained before (P0) and after gastrectomy (P1), (2) intragastric wash samples obtained before any manipulation (G0) and just before resection of the stomach (G1), and (3) ex vivo wash samples obtained by rinsing resected stomach with the lymphovascular pedicles closed by clips (S0) or with the pedicles open (S1). Cytologic examination was performed from all washes, and real-time reverse transcriptase–polymerase chain reaction analysis for carcinoembryonic antigen was performed from washes P0, P1, S0, and S1.ResultsCytologic examination detected cancer cells in 34.2% (13 of 38) of G0 samples and in 39.5% (15 of 38) of G1 samples. The rate of conversion from G0-negative to G1-positive increased as T stage increased. Cytologic examination detected cancer cells in 2.6% (1 of 38) of S0 samples and in 13.2% (5 of 38) of S1 samples. The carcinoembryonic antigen mRNA level of the S1 sample was 2-fold greater than that of the S0 sample in 50.0% (7 of 14).ConclusionsFree cancer cells can be released from gastric lumen or lymphovascular pedicles opened during gastric cancer surgery, especially in advanced-stage disease. Care should be taken to minimize spillage from the gastric lumen and lymphovascular pedicles.

Comprehensive genome- and transcriptome-wide analyses of mutations associated with microsatellite instability in Korean gastric cancers

Microsatellite instability (MSI) is a critical mechanism that drives genetic aberrations in cancer. To identify the entire MS mutation, we performed the first comprehensive genome- and transcriptome-wide analyses of mutations associated with MSI in Korean gastric cancer cell lines and primary tissues. We identified 18,377 MS mutations of five or more repeat nucleotides in coding sequences and untranslated regions of genes, and discovered 139 individual genes whose expression was down-regulated in association with UTR MS mutation. In addition, we found that 90.5% of MS mutations with deletions in gene regions occurred in UTRs. This analysis emphasizes the genetic diversity of MSI-H gastric tumors and provides clues to the mechanistic basis of instability in microsatellite unstable gastric cancers.

Up‐regulated expression of sulfatases (SULF1 and SULF2) as prognostic and metastasis predictive markers in human gastric cancer

Gastric cancer (GC) is the fourth most common cancer worldwide. In spite of the mortality incidence associated with GC, no reliable prognostic biomarkers are currently available for this malignancy. The sulfatases (or SULFs), SULF1 and SULF2, play a critical role in the pathogenesis of a variety of human cancers. We sought to evaluate the potential of SULFs as biomarkers for GC. Thirty pairs of GC and corresponding normal tissues were analysed for the expression and methylation status of SULFs. Furthermore, the functional role of SULF overexpression was investigated in GC cell lines and tumour xenograft animal models. Lastly, we validated the expression of SULF1 protein in a large cohort of 450 GC patients. GC tissues showed conspicuously higher expression of SULF1 (p = 0.0002) and SULF2 (p = 0.001) compared to normal mucosa, which was correlated with its promoter hypomethylation. Furthermore, high expression of SULFs caused marked acceleration in the growth of xenograft tumours in nude mice. The expression of SULF1 protein significantly correlated with higher recurrence rates (p = 0.0002) and worse overall survival (p < 0.0001) in GC patients. Multivariate analysis revealed that SULF1 is an independent prognostic (p = 0.0123) and lymph node metastasis predictive factor (p = 0.0003) in patients with GC. We provide novel evidence that hypomethylation of promoter CpG islands within SULF genes imparts them with oncogenic potential in GC. Moreover, our data suggest that SULF1 may serve as a promising biomarker for patients with GC. Copyright

Injectable and biodegradable poly(organophosphazene) hydrogel as a delivery system of docetaxel for cancer treatment

Abstract Although docetaxel (DTX) is an advanced taxoid, further augmentation of its properties is still required, such as improvement in its low aqueous solubility. Herein, we report the development of biodegradable/injectable poly(organophosphazene) (PPZ) hydrogels for the delivery of DTX without the use of organic solvents. An aqueous solution of PPZ containing α-amino-ω-methoxy-poly(ethylene glycol) (AMPEG) 750 instead of AMPEG 550 was prepared, thereby increasing the erosion capacity of the hydrogel by judicious balance of the hydrophobic/hydrophilic moieties. The safety of the hydrogel was demonstrated using a biocompatibility test. The PPZ aqueous solution (8 wt%) containing DTX exhibited a thermosensitive sol–gel–sol transition that was independent of the concentration of DTX (1–3 mg/mL). The in vitro release study indicated that the dominant release mechanism was either erosion or diffusion/erosion-controlled release depending on the DTX content of the hydrogel. The in vivo anticancer effect of the intratumorally injected PPZ system in human gastric cancer cell-xenografted mice was evaluated, which demonstrated a significantly (p < 0.01) enhanced effect of the DTX-PPZ hydrogel system compared to the control (DTX solution, i.v.). In conclusion, the PPZ hydrogel may be a promising candidate for DTX delivery, affecting a decrease in the size of tumors with little toxicity prior to exeresis.

Overexpression of Plasminogen Activator Inhibitor-1 in Advanced Gastric Cancer with Aggressive Lymph Node Metastasis

Purpose The purpose of this study is to investigate differentially expressed genes using DNA microarray between advanced gastric cancer (AGC) with aggressive lymph node (LN) metastasis and that with a more advanced tumor stage but without LN metastasis. Materials and Methods Five sample pairs of gastric cancer tissue and normal gastric mucosa were taken from three patients with T3N3 stage (highN) and two with T4N0 stage (lowN). Data from triplicate DNA microarray experiments were analyzed, and candidate genes were identified using a volcano plot that showed ≥ 2-fold differential expression and were significant by Welchs t test (p < 0.05) between highN and lowN. Those selected genes were validated independently by reverse-transcriptase–polymerase chain reaction (RT-PCR) using five AGC patients, and tissue-microarray (TMA) comprising 47 AGC patients. Results CFTR, LAMC2, SERPINE2, F2R, MMP7, FN1, TIMP1, plasminogen activator inhibitor-1 (PAI-1), ITGB8, SDS, and TMPRSS4 were commonly up-regulated over 2-fold in highN. REG3A, CD24, ITLN1, and WBP5 were commonly down-regulated over 2-fold in lowN. Among these genes, overexpression of PAI-1 was validated by RT-PCR, and TMA showed 16.7% (7/42) PAI-1 expression in T3N3, but none (0/5) in T4N0 (p=0.393). Conclusion DNA microarray analysis and validation by RT-PCR and TMA showed that overexpression of PAI-1 is related to aggressive LN metastasis in AGC.

Improvement of anti-cancer drug efficacy via thermosensitive hydrogel in peritoneal carcinomatosis in gastric cancer

Peritoneal carcinomatosis (PC) of gastric origin has a poor prognosis with short survival due to lack of effective therapeutic modalities. Here, we evaluated the therapeutic efficacy of an injectable thermosensitive poly (organophosphazene) (PPZ) hydrogel with docetaxel (DTX-gel) to develop an effective therapeutic agent for patient with PC. Three days after inoculation of highly metastatic 44As3Luc cells into peritoneal cavity, the mice were intravenously or intraperitoneally administered with docetaxel alone (DTX-sol IV or IP), and intraperitoneally injected with DTX-gel. The anti-tumor activity was monitored by bioluminescence live imaging system. Compared to DTX-sol IV or IP, the tumor growth was significantly reduced in the DTX-gel treated mice (p<0.0001, p=0.0001). Furthermore, the survival rate was significantly increased in the DTX-gel treated mice compared to DTX-sol IV or IP treated mice (p<0.0001, p=0.0068). Our results demonstrated that DTX-gel suppresses peritoneal metastasis by continuing release of chemotherapy agent, which leads to increase the survival rate in a PC model. Therefore, biodegradable thermosensitive hydrogel with docetaxel system can be a good anti-cancer agent for PC.

Gastric Carcinogenesis in the miR-222/221 Transgenic Mouse Model.

Purpose MicroRNAs (miRNAs) regulate various cellular functions, including development, cell proliferation, apoptosis, and tumorigenesis. Different signatures associated with various tissue types, diagnosis, progression, prognosis, staging, and treatment response have been identified by miRNA expression profiling of human tumors. miRNAs function as oncogenes or as tumor suppressors. The relationship between gastric cancer and miRNA garnered attention due to the high incidence of gastric cancer in Asian countries. miR-222/221 expression increases in gastric tumor tissues. The oncogenic effect of miR-222/221 was previously determined in functional studies and xenograft models. In this study, transgenic mice over-expressing miR-222/221 were generated to confirm the effect of miR-222/221 on gastric carcinogenesis. Materials and Methods At 6 weeks of age, 65 transgenic mice and 53 wild-type mice were given drinking water containing N-nitroso-N-methylurea (MNU) for 5 alternating weeks to induce gastric cancer. The mice were euthanized at 36 weeks of age and histologic analysis was performed. Results Hyperplasia was observed in 3.77% of the wild-type mice and in 18.46% of the transgenic mice (p=0.020). Adenoma was observed in 20.75% of the wild-type mice and 26.15% of the transgenic mice (p=0.522). Carcinoma was observed in 32.08% of the wild-type mice and 41.54% of the transgenic mice (p=0.341). The frequency of hyperplasia, adenoma, and carcinoma was higher in transgenic mice, but the difference was statistically significant only in hyperplasia. Conclusion These results suggest that hyperplasia, a gastric pre-cancerous lesion, is associated with miR-222/221 expression but miR-222/221 expression does not affect tumorigenesis itself.

Abstract 643: Gene methylation as a novel marker in gastric cancer.

Introduction: Epigenetics is one of the major cause of cancer. Several studies have shown the relation between gastric cancer and DNA methylation but it lacks coherence to use as a biomarker. Also there is a lot of genes have not reveled correlation between gastric cancer and DNA methylation. Here, we analyze the correlation between DNA methylation and gene expression in gastric cancer to use as a novel biomarker. Method: Gastric cancer and matched normal gastric tissues were obtained who received gastrectomy in Seoul national university. More than 70 % of tumor share tissues were selected for this study. Nine genes (FAM58A, MAL, TMEM220, MMP28, C2orf3, IL-19, RIN2, HOPX, RGS6) expression level were analyzed using 30 pairs of gastric cancer, normal gastric tissues We then selected genes showing distinct difference mRNA expression level between gastric cancer and gastric tissues to analyze DNA methylation level using bisulfite modification. After that we analyze gene expression level and methylation level using 10 gastric cancer cell line(MKN28, AGS, KATOIII, ect..)to compare with gastric cancer tissues Result: Advanced research of this study performed MIRA for analyze DNA methylation level and Real-time pcr to know mRNA expression level using three snap-frozen gastric cancer and matched normal gastric tissue. This study identified that FAM58A, RYR2, SLC9A7, GHSR, HIST1H4E, MAL, TMEM220,C2orf3, MMP28, EPOR etc sixteen genes were hypermethylated in gastric cancer and there were correlation between methylation level and mRNA expression level Also this study found ten hypomethylated genes (STAC, IL19,HYDIN, C1orf162, OR2J2, RIN2,HOPX, RGS6,MOCS1 ect) using three gastric tissue samples. These genes also associated with mRNA expression level comparison between gastric cancer and normal gastric tissues. Among the hypermethylated genes, we analyzed FAM58A, MAL, TMEM220, MMP28, C2orf3 genes MAL(p mRNA expression level were significantly down-regulated in gastric cancer tissues.. Especially MAL gene is remarkably down-regulated in all 30 samples of gastric cancer tissues. Additionally we analyzed four hypomethylated genes (hand IL-19, RIN2, HOPX, RGS6). RIN2, HOPX, RGS6) but RIN2, HOPX, RGS6 genes mRNA expression level differ in each gastric cancer tissues The other hand IL-19 have shown different mRNA expression level what we expected in advanced study. Except three dramatically up-regulated tissues, almost the others mRNA expression level were down-regulated in 30 gastric cancer tissue samples(p Conclusion: Our finding will provide valuable data for development of the useful gastric cancer methylation marker and understanding of correlation between methylation and mRNA expression level in gastric cancer. Furthermore MAL gene case, we expect it could be a gastric cancer marker not only methylation analysis but also mRNA expression level analysis. Citation Format: Boram Choi, Tae-Su Han, Ji-Yeon Lee, Sunmin Lee, Seong-Ho Kong, Hyuk-Jun Lee, Young-Joon Kim, Han-Kwang Yang. Gene methylation as a novel marker in 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 643. doi:10.1158/1538-7445.AM2013-643