Analysis of Tumor-Related Genes In Pancreatic Juice Aiming to be Useful for Early Diagnosis of Pancreatic Cancer

我们分析了经ERCP抽吸的胰液(PJ)和经PTC或ERBD获得的胆汁DNA提取液(SED)和上清液(SUP)中P53基因的突变。胰腺癌(PCa)患者PJ的Sup和SED中P53基因突变的发生率分别为43%(9/21)和29%(6/21)，与Sup和SED联合检测的P53突变发生率为52%(11/21)。此外，在细胞学诊断为阴性的15例PCa中，有7例(47%)检测到P53突变。在25例慢性胰腺炎(CP)患者中，无一例携带突变型p53。胆道癌患者胆汁中P53基因突变率分别为50%(15/30)和33%(10/30)。另一方面，胆石症患者中没有携带突变型p53的患者。提示sUP中P53突变的发生率高于SED，SUP中P53突变的特异性高于…在SED中，P53突变对癌症的特异性很高。在21例PCa中11例(52%)、11例导管内乳头状黏液性肿瘤(IPMN)中5例(45%)和22例CP中3例(14%)PJ有Mesothlin mRNA表达。PCa组与CP组间皮质醇mRNA表达差异有统计学意义。我们建立了一种高灵敏度的实时定量RT-PCR检测h-TERT mRNA的方法，并检测了血清和PJ中h-TERT mRNA的含量。H-TERT信使核糖核酸水平在几乎所有前列腺癌患者中均升高，但在相当一部分良性病变中也有升高，提示h-TERT信使核糖核酸对肿瘤的特异性较低。良性病变体液中H-TERT m RNA来源于淋巴细胞，应用甲基化特异性聚合酶链式反应分析了PJ组织中ppENK、SARP2和NPTX2 3个CpG岛的甲基化状态。在28例PCa、15例IPMN和20例CP中，PJ中ppENK基因异常甲基化分别为14例(50%)、4例(27%)和1例(15%)，NPTX2基因异常甲基化分别为24例(77%)、16例IPMN中10例(63%)和13例CP中6例(43%)。PCa组ppENK或NPTX2基因异常甲基化发生率明显高于CP组。此外，我们还发现在35例PCa患者中24例(74%)、16例IPMN患者中15例(94%)和19例CP患者中2例(11%)PJ中SARP2基因异常甲基化。在PCa和IPMN中，SARP2基因异常甲基化的发生率明显高于CP。尽管前列腺癌PJ中肿瘤相关基因异常甲基化的发生率非常高，但其对癌症的特异性还不够。IPMN良恶性组之间差异无统计学意义，提示这些基因异常甲基化是PCa发生的早期事件，定量检测PJ异常甲基化可能对PCa具有潜在的诊断意义。较少

We analyzed mutations of p53 in DNA extract not only from the sediment(Sed) but also from the supernatant(sup) of aspirated pancreatic juice(PJ) by ERCP and bile obtained by PTC or ERBD. The incidence of p53 mutations was 43%(9 of 21) and 29%(6 of 21) in the Sup and the Sed of PJ from patients with pancreatic cancer(PCa), respectively, and nurse to 52%(11 of 21) in the combination assay with Sup and Sed. Moreover, p53 mutations were detected in 47%(7 of 15) of PCa cases in which the cyto logic diagnosis was negative. Among 25 patients with chronic pancreatitis(CP), none harbored mutant p53. In bile abtained from patients with biliary tract cancer, the positive rate of p53 mutations was 50%(15 of 30) in the Sup and 33%(10 of 30) in the Sed. On the other hand, there were no pantients with cholelithiasis harboring mutant p53. These results could indicate that the incidence of p53 mutation was higher in the Sup than in the Sed, and the specificity of p53 mutation was higher in the Sup than … More in the Sed, and the specificity of p53 mutation for cancer was very high. Mesothlin mRNA expression in the PJ was discovered in 11(52%) of 21 patients with PCa,5(45%) of 11 with intraductal papillary mucinous neoplasm(IPMN) and 3(14%) of 22 with CP. There was significant difference between PCa and CP for mesothelin mRNA. We developed an highly sensitive quantitative analysis for h-TERT mRNA using real-time RT-PCR and measured the amount of h-TERT mRNA in serum and PJ. The levels of h-TERT mRNA were elevated in almost all cases with PCa but also in considerable cases with benign lesions, suggesting low specificity of h-TERT mRNA for cancer. h-TERT mRNA in body fluids from benign lesions originates from lymphocyte.Using methylation-specific PCR, we nextly analyzed the methylation status of 3 CpG islands of ppENK, SARP2 and NPTX2 in PJ samples. Aberrant methylation of ppENK in the PJ was detected in 14(50%) of 28 patients with PCa, 4(27%) of 15 with IPMN and 1(15%) of 20 with CP, and that of NPTX2 was detected in 24(77%) of 35 patients with PCa, 10(63%) of 16 with IPMN and 6(43%) of 13 with CP. The incidence of aberrant methylation of ppENK or NPTX2 was significantly higher in PCa than in CP. Furthermore, we found aberrant methylation of SARP2 in PJ in 24(74%) of 35 patients with PCa, 15(94%) of 16 with IPMN and 2(11%) of 19 with CP. The incidence of aberrant methylation of SARP2 was significantly higher in PCa or IPMN than in CP. Although the prevalence of aberrant methylation of the tumor-related genes in PJ from PCa was strikingly high, its specificity for cancer was not sufficient. There was no significant difference between benign and malignant groups of IPMN, suggesting that aberrant menthylation of these genes is a early event in PCa development, and quantitative detection of aberrant methylation in PJ may have potential diagnostic implication for PCa. Less