Tin L. Lee

Association between cyclo-oxygenase-2 overexpression and missense p53 mutations in gastric cancer

Wild-type p53 competitively binds to the promoter region of COX-2 in vitro and inhibits its transcription. We examined the association between p53 mutation and COX-2 expression in gastric cancer. COX-2 over-expression was seen in 19 (48.7%) cases. These tumours had more lymph-node metastasis (P = 0.048) and tended to have a poorer survival (P = 0.07). Missense mutations of p53 were detected in 20 (51.3%) patients and had a significantly stronger COX-2 expression than tumours without p53 mutation (P = 0.016). Our results suggest a link between p53 mutation and COX-2 overexpression in gastric cancer.

Hypermethylation of the tumor suppressor gene RASSFIA and frequent concomitant loss of heterozygosity at 3p21 in cervical cancers

Loss of heterozygosity (LOH) at chromosome 3p21 is frequent in cervical cancers. The candidate tumor suppressor gene, RASSF1A located at 3p21.3, is found to be inactivated in several major human cancers, implicating its significance in carcinogenesis. We aimed to investigate the status of RASSF1A in cervical cancers. The mutation and methylation status of RASSF1A were analysed in 4 cervical cancer cell lines, 50 primary cervical cancers including 33 squamous cell carcinoma (SCC), 17 adenocarcinoma (AC) and 11 normal controls. The primary cancer samples were also detected for LOH at 3p21 and human papillomavirus (HPV). Hypermethylation of RASSF1A was detected in 30% of SCC, 12% of AC and in 1 of the 4 cancer cell lines but was absent in all normal cases. Methylation of the cancer cell line was associated with loss of gene expression, which was restored by demethylation. About 67% (8 of 12) of hypermethylated primary cancers showed concomitant LOH at 3p21. No somatic mutation was found in all primary cancer samples or cell lines but 2 cases showed germline polymorphism at codon 133. Oncogenic HPV DNAs were found in most cancer samples. No correlation was detected between RASSF1A‐hypermethylation or LOH at 3p21 and age of patient, HPV genotype, tumor grade and stage. Hypermethylation of RASSF1A occurs in a subset of cervical cancers, among which concomitant LOH at 3p21 is common. The results supported that RASSF1A may be one of the cervical cancer‐related tumor suppressor genes located at 3p21 regions.

Alterations of frizzled (FzE3) and secreted frizzled related protein (hsFRP) expression in gastric cancer

Wnt signaling pathway is important for development and carcinogenesis. Alterations of this pathway, such as mutations in adenomatous polyposis coli (APC) gene and activation mutations of beta-catenin, would result in stabilization of beta-catenin and subsequent translocation to nucleus where genes are transcribed. Recently, a receptor of Wnt, FzE3 was found to be up-regulated in esophageal carcinoma while a non-receptor antagonist of Wnt, secreted frizzled related protein (hsFRP) was found to be down-regulated in some cancer. These findings suggested that FzE3 is a potential oncogene while hsFRP is a potential tumor suppressor gene. We aimed to investigate whether FzE3 and hsFRP were altered in gastric cancer. Twelve cases of gastric cancer, including 7 cases of intestinal type, 4 cases of diffuse type and I case of mixed type, were studied. FzE3 and hsFRP mRNAs were expressed in most of the paired normal gastric tissues. FzE3 was over-expressed in 9 cases (75%) of gastric carcinoma tissues while hsFRP was down-regulated in 2 cases (16%). Beta-catenin nuclear staining was identified in 3 cases (27%) and cyclin D1 was expressed in 5 cases (41%) of cancer samples. All these cases were associated with either up-regulation of FzE3 or down-regulation of hsFRP. Our results suggested that alterations of FzE3 or hsFRP were frequent in gastric cancer. These provide alternative mechanisms leading to activation of Wnt signaling pathway in gastric carcinogenesis.

Expression and cellular localization of COX-1 and -2 in Helicobacter pylori gastritis.

There are conflicting reports on the expression of cyclooxygenase in Helicobacter pylori infection.

Interaction of Helicobacter pylori eradication and non-steroidal anti-inflammatory drugs on gastric epithelial apoptosis and proliferation: implications on ulcerogenesis

Apoptosis is associated with loss of gastric mucosal integrity and may play an important role in ulcer development.

Up-regulation of cyclooxygenase-1 and -2 in human gastric ulcer.

The expression of cyclooxygenase (COX) in human gastric ulcers is unknown.

Somatic β-catenin mutation in gastric carcinoma : an infrequent event that is not specific for microsatellite instability

We screened 90 cases of gastric carcinoma (GCA) samples for beta-catenin exon 3 mutation and assessed its possible relationship with microsatellite instability (MSI). Three mutations were detected in two samples, including a single mutation in an intestinal type and double mutations in a diffuse type GCA. One of the mutations found in the diffuse type GCA sample was a non-sense mutation at codon 68 (CAG-->TAG). This novel mutation was predicted to disrupt the binding of beta-catenin to alpha-catenin and may be related to the diffuse type morphology. The other two mutations were missense mutations involved or related to the GSK-3beta phosphorylation site, which have been reported previously. No MSI can be demonstrated in the two cases with beta-catenin mutation. Our results suggested that beta-catenin mutation was infrequent in GCA and appeared not specific for MSI.

Genome-Wide Binding Analysis of Carboxyl-Terminal Truncated HBx Reveals Direct Repression of a Negative Growth Regulator, GAS2, in Hepatocellular Carcinoma

G A A b st ra ct s disease. TLR2-/-MyD88+/+MDR1A-/macrophages hyperresponded to LPS stimulation with increased production of IL-1β proteinwhich was not evident in TLR2+/+MyD88+/+MDR1A-/macrophages. Conclusions: Exacerbation of UC-like pancolitis in TLR2/MDR1A double deficiency requires IL-1R/MyD88 signalling. Aberrant LPS-induced IL-1β signalling via IL1R/MyD88 may represent a critical inducer of severe pathology and a novel therapeutic target in a subgroup of UC patients.

Abstract 1185: EZH2-mediated H3K27 trimethylation silencing of microRNAs in hepatocellular carcinoma

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Although the biological pathways by which misexpressed microRNAs (miRNAs) contribute to the development of hepatocellular carcinoma (HCC) have been extensively investigated, little is known about the upstream regulatory mechanisms. Epigenetic gene silencing in cancer cells is mediated by promoter DNA methylation and at least two distinct histone modifications, polycomb-based histone H3 lysine 27 trimethylation (H3K27me3) and H3 lysine 9 dimethylation (H3K9me2). We have previously demonstrated that EZH2-mediated H3K27me3 is a mechanism of tumor-suppressor gene silencing in cancer that is potentially independent of promoter DNA methylation (Nat Genet 2008;40:741-50). Here we used an integrative genome-wide location and expression analysis to investigate the epigenetic control of miRNA expression in HCC cells and explored the involvement of transcription factors in this process. Chromatin immunoprecipitation coupled with human promoter microarrays revealed that 8, 20, and 15% of the interrogated miRNA promoters were enriched with H3K27me3, H3K9me2 and DNA methylation in Hep3B HCC cells, respectively. Some of these enriched miRNAs e.g. miR-101 and -194 have been reported to be down-regulated in primary HCCs, providing evidence on epigenetic silencing of miRNAs in HCC. Consistent with our previous observation on protein-coding genes, most of the miRNAs enriched with H3K27me3 had no detectable DNA hypermethylation; and most miRNAs showing DNA hypermethylation had no enrichment for H3K27me3. There was also minimal overlap between H3K27me3 and H3K9me2 binding on miRNA promoters. Down-regulation of EZH2 decreased global H3K27me3 level and restored expression of the H3K27me3-targeted miRNAs while 5-aza-2’-deoxycytidine and trichostatin A treatment alone or in combination did not reactivate their expression, further suggesting the independence of EZH2-mediated H3K27me3 in miRNA silencing. To identify the transcription factors involved in the formation of H3K27me3 modification, the -0.5 to +1kb regulatory regions of ∼2000 H3K27me3-bound protein-coding and miRNA genes were submitted to transcription factor binding predictions by TRANSFAC Pro. Interestingly, the binding sites for YY1, known to recruit polycomb for H3K27me3 modifications, were recurrently over-represented in these loci. Quantitative RT-PCR showed that YY1 and EZH2 were over-expressed (defined as greater than 2-fold increase) in 73% (38/52) and 87% (45/52) of HCCs compared with the paired non-tumor tissues. Overall, their expressions were significantly higher in tumor tissues (p < 0.0001) and positively correlated in these 52 pairs of HCCs (p < 0.05). In conclusion, our findings suggest that YY1 may play an important role in EZH2-mediated H3K27me3 for miRNA silencing in HCC. Acknowledgements: This study was partially supported by the General Research Fund 462309, RFCID 09080042 and Collaborative Research Fund CUHK04/CRF/08. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1185. doi:10.1158/1538-7445.AM2011-1185