Jae Hyuk Lee

CD44 enhances the epithelial-mesenchymal transition in association with colon cancer invasion.

The metastatic process involves the migration and invasion of cancer cells throughout the body to produce secondary tumors at distant sites. Through of epithelial-mesenchymal transition (EMT), cancer cells employ developmental processes to gain migratory and invasive properties. CD44 is the transmembrane adhesion receptor for Hyaluronan (HA) and plays a central role in the remodeling and degradation of HA that leads to cell migration, as well as to cancer invasion and metastasis. CD44 is highly expressed in primary and metastatic colon cancer but lowly expressed in normal tissues. We evaluated the impact of CD44 on EMT and invasion of colon cancer cells. The functional role of CD44 in EMT was determined by the overexpression or knockdown of CD44. CD44 was overexpressed by transfection with plasmid-RT-PCR product and knockdown of CD44 by small hairpin RNA (shRNA)-mediated depletion of CD44 in SW480 colon cancer cells. Morphological changes were evaluated by confocal laser microscopy in the culture media. The expression of EMT markers (E-cadherin/N-cadherin/vimentin/fibronectin/actin/MMPs) and CD44/EGFR/PI3K-Akt signaling were evaluated using western blotting. The influence of EMT in tumor biology was assessed with proliferation, migration and invasion assays. EMT changes increased in CD44-overexpressing SW480 cells and decreased in CD44 knockdown cells. CD44 activation induced expression of EGFR and activation of phosphatidylinositol 3 kinase (PI3K)/Akt and expression of glycogen synthase kinase-3 β (GSK-3β). In terms of EMT markers, CD44 downregulated E-cadherin expression, upregulated N-cadherin, α-actin, vimentin, fibronectin and MT1-MMP, and inhibited the formation of the membrane-associated E-cadherin-β-catenin complex, which resulted in cell invasion and migration.

TMPRSS4 induces invasion and epithelial–mesenchymal transition through upregulation of integrin α5 and its signaling pathways

TMPRSS4 is a novel type II transmembrane serine protease that is highly expressed on the cell surface in pancreatic, thyroid and other cancer tissues, although its oncogenic significance and molecular mechanisms are unknown. Previously, we have shown that TMPRSS4 promotes invasion, migration and metastasis of human tumor cells by facilitating an epithelial-mesenchymal transition (EMT). In this study, we explored the molecular basis underlying TMPRSS4-mediated effects. We show that multiple downstream signaling pathways, including focal adhesion kinase (FAK), extracellular signal-regulated kinase (ERK), Akt, Src and Rac1, are activated by TMPRSS4 expression and that FAK signaling and ERK activation are required for TMPRSS4-induced invasiveness and EMT, including cadherin switch. Inhibition of PI3K or Src reduced invasiveness and actin rearrangement mediated by TMPRSS4 without restoring E-cadherin expression. Downregulation of E-cadherin was required for TMPRSS4-mediated effects but was not sufficient to induce EMT and invasion. TMPRSS4 induced integrin alpha5 expression and its signal transduction, leading to invasiveness and EMT accompanied by downregulation of E-cadherin. Functional blocking confirmed that integrin alpha5beta1 is a critical signaling molecule that is sufficient to induce TMPRSS4-mediated effects. Immunohistochemical analysis showed that TMPRSS4 expression was significantly higher in human colorectal cancer tissues from advanced stages than in that of early stage. Furthermore, upregulation of TMPRSS4 was correlated with enhanced integrin alpha5 expression. These observations implicate integrin alpha5 upregulation as a molecular mechanism by which TMPRSS4 induces invasion and contributes to cancer progression.

BRCA1 and XRCC1 polymorphisms associated with survival in advanced gastric cancer treated with taxane and cisplatin

This study evaluated the influence of genetic polymorphism influencing drug metabolism on survival in taxane‐ and cisplatin‐treated advanced gastric cancer (AGC). Peripheral blood samples from 207 AGC patients treated with first‐line chemotherapy of taxane and cisplatin were used. We investigated polymorphisms that influenced the metabolism of taxane (ATP‐binding cassette transporter B1 (ABCB1)), cisplatin (glutathione S‐transferase M1 (GSTM1), glutathione S‐transferase P1 (GSTP1), glutathione S‐transferase T1 (GSTT1), excision repair cross complementing 1 (ERCC1), X‐ray Cross Complementing group 3 (XRCC3), X‐ray Cross Complementing group 4 (XRCC4), X‐ray Cross Complementing group 1 (XRCC1), breast cancer (BRCA1)), and 5‐fluorouracil (methylene tetrahydrofolate reductase (MTHFR), thymidylate synthase (TYMS)). A total of 207 patients were enrolled between May 2004 and Dec 2008, and 200 patients were analyzed. The overall response rate was 38.5%. Time to progression and overall survival time were 4.3u2003±u20030.19u2003months and 11.9u2003±u20031.05u2003months, respectively. There was no significant association between genetic polymorphism and response rate. However, the BRCA1 mutant TT homozygote was associated with significant prolongation of overall survival (hazard ratio [HR]u2003=u20030.43; 95% confidence interval [CI], 0.20–0.92; Pu2003=u20030.03) and progression‐free survival (HRu2003=u20030.51; 95% CI, 0.26–1.00; Pu2003=u20030.05). Also, the XRCC1 194 CT genotype was associated with inferior overall survival, relative to the XRCC1 194 CC homozygotes (HRu2003=u20031.49; 95% CI, 0.11–2.07; Pu2003=u20030.018).These findings suggest that BRCA1 TT and XRCC1 194 CT genotypes could be modest prognostic markers of AGC response in taxane‐ and cisplatin‐treated patients.

Expression of standard CD44 in human colorectal carcinoma: association with prognosis.

The aim of the present study was to evaluate the expression of standard CD44 (CD44s) in colorectal cancer (CRC), its relationship with clinicopathological characteristics, and its potential prognostic significance. CD44s levels were measured on immunohistochemistry in tumors and surrounding normal mucosa from 74 patients with primary colorectal carcinomas. The patients were followed for a median period of 37u2003months. Expression of CD44s in primary tumor and surrounding normal mucosa tissues was demonstrated in 100% (74/74) and 37.9% (28/74), respectively. The expression of CD44s in tumors was significantly associated with the depth of invasion (Pu2003=u20030.034) and lymph node involvement (Pu2003=u20030.031). A significant difference was observed between the overall survival and level of tumor CD44s expression, especially for stage IV carcinoma (Pu2003=u20030.038). Multivariate analysis indicated that TNM stage (Pu2003=u20030.020) and tumor CD44s expression (Pu2003=u20030.008) were independent predictors of overall survival in adenocarcinomas. CD44s overexpression may be an independent unfavorable prognostic factor for overall survival in advanced CRC, especially stage IV disease. Further investigation, however, is necessary to assess the biological roles of CD44 in CRC, and validate their possible value as novel therapeutic targets.

Aberrant methylation of DNA mismatch repair genes in elderly patients with sporadic gastric carcinoma: A comparison with younger patients.

Hypermethylation of promoters that regulate the expression of DNA repair genes is associated with gastric carcinoma (GC). Little is known regarding the association between age of disease onset and differences in molecular profiles.

Immunocytochemical panel for distinguishing between adenocarcinomas and reactive mesothelial cells in effusion cell blocks.

The aim of our study was to determine the value of a panel that consisted of one epithelial marker (MOC‐31) and two mesothelial markers (D2‐40 and calretinin) for distinguishing between reactive mesothelial cells (RMCs) and adenocarcinomas (ACs) in effusion fluids. A total of 118 cell block specimens from pleural and peritoneal effusions, including 88 ACs and 30 benign effusions with RMCs were stained with antibodies against MOC‐31, D2‐40, and calretinin. MOC‐31 membranous activity was observed in all samples from ACs, regardless of the primary tumor site. All benign effusion samples with RMCs were negative for MOC‐31. All benign effusion samples with RMCs exhibited membranous staining for D2‐40, and one AC case had focal reactivity for D2‐40. Almost all benign effusions reacted positively with calretinin. Staining was noted in both the cytoplasm and the nucleus in the majority of cases. Scattered tumor cells had weak calretinin positivity in two AC cases. Background RMCs in AC effusions were consistently positive for D2‐40 and calretinin. In general, D2‐40 identified more RMCs than calretinin. The staining combination of positive for MOC‐31 and negative for D2‐40 or calretinin were 100% specific and 99% sensitive for ACs. Our data suggest that immunohistochemical studies performed on cell blocks with MOC‐31, D2‐40, and calretinin were useful in the differentiation between ACs and RMCs. D2‐40 was a more sensitive marker for RMCs than calretinin. Diagn. Cytopathol. 2009.

Quantitative promoter hypermethylation profiles of ductal carcinoma in situ in North American and Korean women: Potential applications for diagnosis

To investigate the diagnostic potential of DNA methylation-based markers in tissuesamples of DCIS, we examined the prevalence and extent of methylation in breastductal carcinoma in situ (DCIS) samples from North American and Korean women.Quantitative multiplex-methylation specific PCR (QM-MSP) of ten genes wasperformed. The methylation level of APC1, Cyclin D2, HIN-1, RASSF1A, and Twistsingly, and cumulative methylation of all ten genes was significantly higher in DCIScompared to normal tissues for both groups. A three-gene panel (APC1, HIN-1, andRASSF1A), QM-MSP distinguished between DCIS and normal breast tissues with asensitivity of 94 to 96% and a specificity of 81 to 87 %. Methylation levels of thesethree genes in DCIS were higher than those of hyperplasia or adjacent normal appearingtissues in Korean women. Comparing North American and Korean DCIS, statisticallysignificant differences in methylation levels were found for CDH1, ER α and RAR- β.Quantification of gene methylation combined with immunohistochemistry in a smallsubset of tumors suggested that methylation may precede loss of protein expression forER-α. Our study demonstrated that methylation profiles of DCIS between NorthAmerican and Korean women were similar. Methylation status of a panel of genesmeasured in a quantitative manner accurately discriminated between normal and DCIStissues of both groups. For both North American and Korean women, QM-MSPanalysis of a key panel of genes may be useful as an ancillary tool for DCIS detection inbreast tissues.

Basal-like breast cancer displays distinct patterns of promoter methylation

Recent microarray profiling studies on breast cancer have identified distinct subtypes that are associated with different clinical outcomes. Promoter hypermethylation of several known or putative tumor suppressor genes occurs frequently during the pathogenesis of breast cancer. We proposed that immunohistopathologic subtypes of breast cancer are likely to contain distinct promoter methylation patterns. A panel of 10 gene promoters was assessed by quantitative multiplex methylation-specific PCR in 114 invasive ductal carcinomas from Korea representing the three major subtypes [57 luminal, 24 human epidermal growth factor 2 (HER2), and 33 basal-like] based on immunohistochemical findings of estrogen receptor, progesterone receptor, HER2, cytokeratin 5/6, and epidermal growth factor receptor.The median methylation levels of HIN1, RASSF1A, and TWIST, and the average methylation ratio were significantly lower in basal-like subtype compared to luminal or HER2 subtypes. In contrast, BRCA1 methylation level was significantly higher in basal-like subtype than in luminal subtype. The methylation status of a panel of four genes (APC1, CDH, BRCA1 and RAR-b) inluminal and HER2 subtypes were dissimilar, where HER2 tumors showed a significantly higher level of methylation compared to luminal tumors. These results suggest that gene methylation in breast cancer can potentially serve as epigenetic biomarkers and may contribute further to current breast cancer classification.

E-cadherin expression in early gastric carcinoma and correlation with lymph node metastasis.

Abnormal expression of E‐cadherin plays an important role in the differentiation and progression of gastric carcinoma. However, the relationship between molecular changes in E‐cadherin and metastasis in early gastric carcinoma (EGC) is poorly understood.

CD133 mRNA expression and microsatellite instability in colorectal carcinoma.

The present study was performed to examine the CD133 expression in colorectal cancer and to analyze its relationship with microsatellite instability (MSI) and the clinicopathological factors, including patient survival.