Whole Genome Exon Sequencing And Clinicopathological Study Of Pancreatic Solid Pseudopapillary Neoplasm

Pancreatic solid pseudopapillary neoplasm (SPN) is a rare tumor of pancreatic with malignant potential. The incidence of pancreatic SPN is very low, accounting for only 1%-2% of all pancreatic tumors, and usually found in young women. The onset of tumor was insidious, as most patients had no clinical symptoms. Patients with symptoms showed early satiety, abdominal discomfort, nausea, vomiting, tumors located in the head of pancreas occasionally showed jaundice. Because of the growing awareness and improved imaging techniques, the detection rate of SPN is gradually increased. The pancreatic SPN was a rare tumor with unique clinical features and biological behavior, but the relevant experimental and clinical research was little.Gene mutation has been widely used in research related to cancer. It has played an important role in reveal the development process of tumor. The next-generation sequencing technology (NGS) is characterized by high-throughput sequencing, including whole genome sequencing and exon sequencing. Exon sequencing is the use of DNA sequencing by NGS technology and target sequence capture technology to capture all of the exon region of the genome. The sequencing process was "cycle sequencing chip method", which was based on repeated reading of the DNA polymerase reaction (template denaturation, primer annealing and extension hybridization) and fluorescence sequence to chips full of DNA samples. Exon sequencing was a targeted, deep coverage, high accuracy technology, with the advantage of simple, economical, efficient and so on, had been widely used in the study of tumor diseases.In this study, we extracted DNA from the fresh tissue samples of pancreatic SPN, constructed DNA libraries and went through high throughput exon sequencing, and screened 94 gene mutations. Further mutations bioinformatic analysis and Sanger sequencing confirmed CTNNB1 as the significant mutations of pancreatic SPN. The expand PCR validation of the 12 pancreatic SPN tumor tissues showed that CTNNB1 mutation was 93.3%(14/15). For a total of CTNNB1 encoding p-catenin protein in the 91 cases of immunohistochemical staining of paraffin tissue sections were stained in 87 cases, the results show a total of 72 cases of tissue sections were positive, and the positive rate was 82.8%(72/87). Correlation analysis showed that the immunohistochemical expression of β-catenin positive degree and tumor diameter was correlated (P=0.029).All patients with complete clinicopathological records who underwent surgery for SPN between 2000 and 2010 were retrospective reviewed.Of the 100 patients identified,84 (84.0%) were female, and the median age was 31 (range,13-68) years old. Median tumor size was 6.5 (range,1.5-18) cm. Twenty-four patients (24.0%) were classified to have malignant SPN. Forty-nine patients had lymph node removed in surgery, and four (8.2%) had nodal metastases. On univariate analysis, peri-pancreatic lymphadenopathy on preoperative computed tomography (CT) and/or magnetic resonance (MR) images was significant risk factor of malignancy (P= 0.025). In the long-term follow up, two patients had evidence of liver metastases and underwent a second laparotomy for metastatic tumor. These two patients were followed up for 24 and 32 months respectively, and never presented with tumor recurrence again.In summary, this thesis conducted a series of experiments on the molecular mechanisms and clinical pathological features of pancreatic SPN. The results showed that mutation of CTNNB1 gene played an important role in the happen and development of pancreatic SPN. The expression of β-catenin was closely related to the tumor proliferation of pancreatic SPN. This gene is hope to become a diagnostic and therapeutic molecular target of pancreatic SPN in the future. Furthermore, this study proved that peripancreatic lymphadenopathy on preoperative radiologic images was associated with malignancy in patients with pancreatic SPN. The long-term follow-up suggested that close follow-up and review periodically were recommended for patients with malignant pancreatic SPN.