Retention of 18q alleles in microsatellite-stable cancers and mutation of the gene for the type II receptor for TGF-beta1 in cancers with high levels of microsatellite instability point to a favorable outcome after adjuvant chemotherapy with fluorouracil-based regimens for stage III colon cancer.

Intraductal papillary-mucinous neoplasms (IPMNs) of the pancreas are a distinct clinicopathological entity, characterized by dilated pancreatic ducts and ductules that are lined by papillary proliferations of tall columnar mucin-producing neoplastic epithelial cells.

, we found that S100A4, a calcium-binding protein previously implicated in metastasis, was expressed in five of seven pancreatic carcinoma libraries but not in the two normal pancreatic duct libraries. We confirmed the overexpression of S100A4 using reverse transcriptase-polymerase chain reaction, which demonstrated that 18 of 19 (95%) pancreatic carcinoma cell lines expressed S100A4. Using immunohistochemistry, we found that 57 of 61 invasive pancreatic carcinomas (93%), 3 of 18 high-grade pancreatic intraepithelial neoplasia lesions (17%), and 0 of the 69 low-grade pancreatic intraepithelial neoplasia lesions expressed S100A4 protein, whereas normal pancreatic tissue and tissue affected by chronic pancreatitis did not label. Expression of S100A4 was associated with poor differentiation of the pancreatic adenocarcinomas (P ‫؍‬ 0.001). We found that three CpG sites in the first intron of the S100A4 gene were ϳ90% methylated in microdissected normal pancreatic duct cells using bisulfite-modified sequencing and in two cell lines and three primary pancreatic carcinomas with a reduced or absent expression of S100A4. In contrast, these CpGs were 100% hypomethylated in 11 of 12 pancreatic cancer cell lines by methylation-specific polymerase chain reaction. The association between the expression of S100A4 and hypomethylation of the first intron of S100A4 was statistically significant (P ‫؍‬ 0.002). These data suggest that the majority of pancreatic carcinomas undergo selection for hypomethylation and overexpression of S100A4. Because most pancreatic carcinomas express S100A4, it may be a useful target for early detection strategies. (Am J Pathol 2002, 160:45-50) S100A4 (also called mts1, p9Ka, calvasculin, CAPL, pEL98) is a member of a family of sixteen S100 calciumbinding proteins, that all have in common a functional EF-hand domain that mediates their activity.1 The S100A4 gene was originally cloned by differential screening experiments in which cDNAs were compared between cells before and after growth stimulation or transformation by oncogenes.2-4 S100A4 is thought to promote metastasis. Nonmetastatic tumor cell lines transfected with S100A4 have a higher incidence of metastases and increased motility.5 S100A4 transcripts are increased in tumor rat cell lines with metastatic properties compared to their nonmetastatic counterparts. 6 The mechanisms by which S100A4 is overexpressed in cancer cell lines has been studied and hypomethylation of CpG sites in the S100A4 gene has been associated with overexpression in colorectal and lymphoma carcinoma cell lines. 7,8 In an initial survey of gene expression analysis of pancreatic cancer using serial analysis of gene expression (SAGE), we identified 47 tags that were overexpressed in pancreatic cancer cell lines compared to normal pancreatic ductal cells (Ryu B, Jones J, Hollingsworth MA, Hruban RH, Kern SG, submitted).9 Among these tags, one corresponded to S100A4 mRNA. Two-thousand threehundred and nine S100A4 tags per million were found in the pancreatic cancer cell lines ...

Pancreatic intraductal neoplasia (PanIN) is thought to be the precursor to infiltrating pancreatic ductal adenocarcinoma. We have previously shown that the preproenkephalin (ppENK) and p16 genes are aberrantly methylated in pancreatic adenocarcinoma. In this study we define the methylation status of the ppENK and p16 genes in various grades of PanINs. One hundred seventy-four samples (28 nonneoplastic pancreatic epithelia, 7 reactive epithelia, 29 PanIN-1A, 48 PanIN-1B, 27 PanIN-2, 14 PanIN-3, 15 invasive ductal adenocarcinomas, and 6 miscellaneous pancreatic neoplasms) were microdissected from 29 formalin-fixed paraffin-embedded surgically resected pancreata, and were analyzed by methylation-specific polymerase chain reaction. Intraductal lesions are commonly found in the pancreatic ducts adjacent to invasive ductal adenocarcinoma, and have been regarded as precursor lesions of pancreatic ductal adenocarcinoma.1-3 Recently, pancreatic ductal lesions, previously described as hyperplasias or dysplasias, have been reclassified into four groups of pancreatic intraepithelial neoplasias (PanINs): PanIN-1A, -1B, -2, and -3.2,3 Molecular analysis of PanIN lesions could help refine the PanIN classification and provide insights into molecular events important to the pathogenesis of early pancreatic ductal adenocarcinoma.The K-ras oncogene is frequently mutated in pancreatic carcinomas as are a number of tumor suppressor genes such as p53, p16, DPC4/SMAD4, 4 -8 and less often BRCA2, TGF␤R1, TGF␤R2, BRCA2, ALK4, STK11, Some PanINs harbor the same genetic changes seen in invasive ductal adenocarcinomas, albeit at lower frequency. 4 -9 Mutations of the K-ras and p16 genes are occasionally found in low-grade PanINs but are observed more frequently in high-grade PanINs. -6SMAD4 inactivation is observed at the PanIN-3 stage.

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Background. Gastric carcinoma with lymphoid stroma (GCLS) has been reported to have a more favorable prognosis than ordinary gastric carcinoma, however, the precise mechanism of the pathogenesis for GCLS remains unclear. Methods. The authors analyzed 99 GCLS in 94 patients for Epstein‐Barr virus (EBV) sequences using polymerase chain reaction (PCR) and in situ hybridization (ISH); these were compared with 42 ordinary gastric carcinomas. Results. Two series of PCR showed 81 (82%) and 46 (47%) of the 99 GCLS to have EBV sequences, which were significantly higher compared with ordinary gastric carcinoma (50% and 9.5%, respectively). With ISH using thymine‐thymine dimerized oligonucleotide probes corresponding to EBV‐encoded small RNA 1 (EBER1), 82 (83%) of 99 GCLS showed clear, intense hybridization signals localized over the nuclei of the tumor cells, in contrast to only 4 (9.5%) of 42 ordinary carcinomas (P < 0.001). A comparative morphologic analysis of EBER1‐positive and negative GCLS revealed that typical features of GCLS, such as mild cellular pleomorphism, rare mitoses, a marked degree of lymphoid stroma, and mild fibrosis, together with a lymphoid infiltration within the cancer cell nests were significantly more frequent in EBER1‐positive GCLS. Conclusions. More than 80% of GCLS were associated with EBV. The presence of EBV association in GCLS was characterized by the above morphologic features.

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The aim of this study was to determine the utility of detecting methylated ppENK and pi6 in pancreatic juice by methylation specific PCR as a marker of pancreatic adeno-carcinoma. Pancreatic juice samples were collected either intraoperatively, from 92 patients undergoing pancreaticoduodenectomy for benign (n=20) and malignant periampullary disease (n = 72) or endoscopically (by duodenal aspiration after secretin infusion), from 13 patients undergoing investigation for pancreatic disease. Methylated ppENK was detected in the pancreatic juice of 30 (66.7%) of 45 patients with pancreatic ductal adenocarcinoma, in 4 (44.4%) of 9 patients with intraductal papillary-mucinous adenocarcinoma, and in 7 (41.2%) of 17 patients with other periampullary carcinomas, using methylation specific PCR. Methylated pi6 was detected in a lower percentage of these patients (11.1%, 11.1% and 23.5%, respectively). In contrast, methylated ppENK and pi6 were not detected in 20 patients with non-malignant periampullary disease including 12 patients with chronic pancreatitis. Methylated ppENK was detected in 30 of 33 (90.9%) primary pancreatic adenocarcinoma and methylated pi6 was in 6/33 (1 8.2%). Despite the absence of ppENKand pi6 methylation in normal pancreas, methylated ppENK and pi6 was present in the duodenum of 90.5% and 28.6%, respectively of patients without cancer. Further, methylated ppENK and pi6 was seen in 88.9% and 11.1%, respectively of pancreatic juice samples obtained by duodenal aspiration from patients without cancer. We conclude that since ppENK and pi6 are not normally methylated in pancreatic secretions, detection of methylated ppENK and pi6 in pure pancreatic juice obtained by direct cannulation of the pancreatic duct to avoid duodenal secretions may suggest the presence of pancreatic adenocarcinoma