| Research Abstract |
In order to identify bradn-new methylation associated molecules, we performed pharmacological unmasking microarray assay subsequent to the addition of inhibitory agents for deacetylation as well as the demethylating agent. Simultaneously, we performed Laser Microdissection assay to identify cancer cell specific repressed genes in comparison to the corresponding norIn order to identify bradn-new methylation associated molecules, we performed pharmacological unmasking microarray assay subsequent to the addition of inhibitory agents for deacetylation as well as the demethylating agent. Simultaneously, we performed Laser Microdissection assay to identify cancer cell specific repressed genes in comparison to the corresponding normal counter part. The microarray analysis (Agilent Technology, 12,814 genes) was performed with gastricd cancer cells extracted specifically by LMD. Therefore, we obtained overlapped gene expression profiles between methylated genes and cancer cell respressed genes.
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Then, this combined method enabled us to uncover 34 genes those might be occult cancer suppressor genes.
Among identified 34 genes, we focused on Fhll gene. By real time quantitative RTPCR assay and immunohistochemical analysis, Fhll expression was significantly downregulated in gastric cancer cells. This result was consistent with the previous reports that Fhll expression was suppressed in various types of malignancies. Concerning regulation mechanism of Fhll expression, Shen Y., et. al. identified Fhll gene as downstream of Crk-associated substrate (Cas) signaling network. According to their reports, Fhll expression appears to be down regulation by phosphorylated Cas in this network. Additionally, many studies have indicated that Cas signaling network promotes cancer development. Among them, Brabek J., et. al. demonstrated that activation of Cas network drove cancer cells to increase their capacity of invasiveness by in vivo assay. Taken together with the evidence that Fhll gene acts cancer suppressor effects by maintaining intercellular junction, it is suggested that Cas signaling network blocks Fhll to achieve its role, resulting in the collapse of intercellular junction. Loss of cell-to-cell communication is the first step for tumor progression. In fact, our survival curves plotted by Kaplan-Meier method demonstrated that patients with all lower expression tumor showed shorter survive, due to deeper tumor invasion or more frequent distant metastasis: Reduced expression of Fhll may be a molecular trigger to aecelerate tumor invasion and migration through Cas signaling network.
The most intriguing aspect of this study is positive correlation between Fhll lower expression and the incidence of distant metastasis. It is indeed that multivariate analysis demonstrated that Fhll lower expression was not an independent indicator for distant metastasis. However, loss of Fhll expression was more closely correlated with distant metastasis. As a matter of fact, there are several reports concerning inverse relationshio between Fhl1 expression and distant metastasis. For instance, La Tulippe E., et. al. reported that the result of gene expression profiling comparing primary prostate cancers with metastatic prostate cancers using microarray analysis demonstrated that Fhl1 was under expressed in metastatic cancers. Additionally, it was reported that Fhl1 expression in metastatic site was down-regulated compared with that in the matched normal mucosa regarding breast cancer and malanoma. Taken these above reports into consideration, it is suggested that loss of Fhl1 expression in cancer cells play a crucial role in acquisition of metastatic propensites
Our serial study revealed the evidence that of Fhl1 expression in primary site of gastric cancer, not metastatic site, may be more reliable indicator for distant metastatis than venous permeation. Because the incidence of distant metastasis is the most influential factor for clinical outcome, it is important for the manaferment of gastric cancer patients to detect the presence of distant metastasis with accuracy. However, it is difficult to estimate increased ability of distant metastasis from pathologically venous permeation evaluated on sections routinely stained with hematoxylin-eosin, and Fhl1 lower expression in primary site may be a powerful diagnostic modality to predict metastatic propensity of gastric cancer cells more easily and accurately.
In conclusion, this current study revealed that clinicopathologic significances of Fhl1 expression in gastric cancer. Fhl1 expression was down-regulated in gastric cancer, ant the patients with Fhl1 lower expression tumor showed survive due to inverse relationship between Ghl1 expression and poor prognostic factor such as deeper tumor invasion and more frequent distant metastasis. Although Fhl1 lower expression was not an independent indicator for distant metastasis by multivariate analysis, it was more significantly correlated with distant metastasis than venous permission. Loss of expression Fhl1 expression may be a potencial indicator for distant metastasis in gastric cancer. Less
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