| Pancreatic cancer, a fatal malignant tumor, is difficult to diagnose, and it lacks of effective treatment. There is an urgent call for effective biomarkers for diagnosis and curative targeted drugs in clinical practice. DNA methylation plays an important role in its pathologic process. The study of gene methylation may become a breakthrough in the diagnosis and treatment of this disease. Illumina 450K Infinium Methylation BeadChip was used to detect pancreatic carcinoma tissue (B), adjacent tissue (C) and corresponding blood samples(A). BeadChip scanning and data analysis were applied. Meanwhile, representative genes were verified to find effective diagnostic markers and new therapeutic targets.The first part:Differentially methylated CpG sites of pancreatic cancer tissues and corresponding adjacent tissues were compared to explicit genome-wide methylation profiles. Results:l.we observed good linearly relationship of genome methylated sites between pancreatic cancer tissue and paracancerous tissue.2. The number of genome-wide methylated CpG sites were less than unmethylated CpG.3.We obtained important signaling pathways and molecular functions as well as biological processes and cellular components.4.We further analyze the high frequent methylatd genes involved in KEGG GO and signaling Pathway. 5.We obtained hypermethylated gene and hypomethylated gene in promoter area by analyzing the level of methylated CpG sites.6. We obtained the cancer signaling pathway based on the differential methylated genes.The second part:We have compared the differentially methylated gene of (CvsB) and (AvsB) to obtain the overlapped differentially methylated gene set (overlap set), which was considered as the common differentially methylated CpG sites of cancer tissues and blood samples. The analysis of overlap set confirmed the blood DNA methylation profiling and patterns. Results:1.differentially methylated sites in blood samples of pancreatic patients and paracancerous tissue showed good linear relationship.2. Cluster analysis showed the difference among groups (A, B and C) was significant, while no significantly difference within each group was observed, and the results reflected the representative of research objects.3. In overlap set, the differential methylation CpG sites genome location were revealed.4. The level and frequency of blood differentially methylated genes were obtained.5. we extracted the methylated sites of promoter region, sorted the data in accordance with level of methylation and clarified the modified model of differentially methylated genes.6. GO and Pathway analysis of important differential methylated genes in promoter region were performed.7. KEGG signaling pathway analysis of pancreatic cancer indicated important differentially methylated genes in promoter region, which was involved in pancreatic carcinogenesis and development.The third part:We take the modified model of serum differentially methylated genes into consideration as well as related studies reported. We selected MAD1L1 for verification by flight mass spectrometry to identify the feasibility of MAD1L1 as diagnostic markers of pancreatic cancer.The fourth part:We take the Beadchip results into consideration in addition to the related results concerning ASS1 reported in the literature. Consequently, we selected ASS1 as research target. ASS1 is a key gene which regulates arginine synthesis and we confirmed the different ASS1 methylation level between cell lines. We also clarified the effect of arginine acid deprivation on ASS1 expression and the mechanism of methylation inhibitor 5-AZA. Finally, we discussed the mechanism that the arginine deprivation inhibited pancreatic cancer cell migration, invasion and clustering, which was the theoretical foundation for the "hunger" treatment of pancreatic cancer. |
PDF Full Text Request