Patrick D. J. Sturm

K-ras oncogene mutations in osteoclast-like giant cell tumors of the pancreas and liver: genetic evidence to support origin from the duct epithelium.

Osteoclast-like giant cell tumors (OCGTs) of the pancreas and liver are enigmatic tumors. Despite their striking morphologic resemblance to certain mesenchymal tumors of bone and tendon sheath, it has been suggested that these tumors may, in fact, arise from epithelial precursors. It is also unclear whether the osteoclast-like giant cells in OCGTs are neoplastic or nonneoplastic. We identified OCGTs of the pancreas and liver that were associated with atypical intraductal epithelial proliferations or mucinous cystic neoplasms. To determine the relationship between the noninvasive epithelial proliferations and the infiltrating OCGTs, each individual component was analyzed for mutations at codon 12 of the K-ras oncogene. Four of the five-duct epithelial lesions harbored activating mutations of the K-ras oncogene. In each case, the same K-ras mutation was also present in the mononuclear cells from the paired OCGT. Moreover, these same mutations were detected when the osteoclast-like giant cells were individually microdissected and analyzed. A panel of immunohistochemical stains was performed, and the osteoclast-like giant cells demonstrated macrophage differentiation. These cells were consistently reactive for the monocyte/macrophage marker KP1, but showed absent staining for a panel of epithelial markers. The infiltrating mononuclear cells lacked strong staining for epithelial markers and monocyte/macrophage markers. These findings suggest that OCGTs of the pancreas and liver are undifferentiated carcinomas that arise directly from intraductal epithelial precursors. The finding of K-ras mutations in the osteoclast-like giant cells may reflect their propensity to phagocytize tumor cells.

The potential diagnostic use of K-ras codon 12 and p53 alterations in brush cytology from the pancreatic head region

It can be difficult to distinguish between malignant and benign disease of the region of the head of the pancreas using conventional methods. K‐ras and p53 alterations occur frequently in malignancies in this region and are therefore candidate tumour markers. To define the utility of these alterations in interpreting pancreatic head cytology, the present study investigated to what extent alterations in the carcinomas were detectable on cytology and whether the alterations found in the cytology came from the carcinomas. Fifty‐seven consecutive pancreaticoduodenectomy resection specimens (52 with a malignancy and five without) and the ductal brush cytology specimens collected post‐operatively from these resection specimens were compared for the presence of K‐ras and p53 alterations. K‐ras mutations were detected using the polymerase chain reaction (PCR) and allele‐specific oligonucleotide hybridization, and p53 alterations using immunochemical staining for the p53 gene product. After discrepancy analysis, the results from the resection specimens and corresponding brush cytology specimens were identical in 88 per cent for the K‐ras analysis and in 84 per cent for the p53 analysis. In two cases, K‐ras mutations found in the brush cytology specimens were not derived from the carcinoma but from pancreatic ductal hyperplasias. Intratumour heterogeneity and sampling error were also identified as causes for discrepant results. The five resection specimens without a malignancy and the corresponding brush cytology specimens were negative for both genetic alterations. In conclusion, the detection of K‐ras and p53 alterations in cells obtained from the pancreatic head region might be a valuable adjunct to conventional cytology for the diagnosis of malignancies in the pancreatic head region. However, intratumour heterogeneity, mucinous pancreatic duct hyperplasia harbouring K‐ras mutations, and sampling error will hinder their diagnostic accuracy in routine clinical use.

Diagnostic and prognostic value of incidence of k-ras codon 12 mutations in resected distal bile duct carcinoma

Background and Objectives: The K‐ras gene is one of the most extensively investigated oncogenes in a wide variety of human tumors, but has rarely been studied in distal bile duct carcinoma (DBDC). We sought to investigate the diagnostic and prognostic value of K‐ras codon 12 mutations in this type of tumor.

Cytology of peritoneal lavage performed during staging laparoscopy for gastrointestinal malignancies: is it useful?

OBJECTIVE To evaluate the potential benefit of cytology of the peritoneal lavage obtained during diagnostic laparoscopy for staging gastrointestinal (GI) malignancies. SUMMARY BACKGROUND DATA Peritoneal lavage is a simple procedure that can be performed during laparotomy for GI tumors. Tumor cells in the lavage fluid are thought to indicate intraperitoneal tumor seeding and to have a negative effect on survival. For this reason, peritoneal lavage is frequently added to diagnostic laparoscopy for staging GI malignancies. METHODS Patients who underwent peritoneal lavage during laparoscopic staging for GI malignancies between June 1992 and September 1997 were included. Lavage fluids were stained using Giemsa and Papanicolaou methods. Cytology results were correlated with the presence of metastases and tumor ingrowth found during laparoscopy and with survival. RESULTS Cytology of peritoneal lavage was performed in 449 patients. Tumor cells were found in 28 patients (6%): 8/87 with an esophageal tumor, 2/32 with liver metastases, 11/72 with a proximal bile duct tumor, 7/236 with a periampullary tumor, and none in 7 and 15 patients with a primary liver tumor or pancreatic body or tail tumor, respectively. In 19 of the 28 patients (68%) in whom tumor cells were found, metastatic disease was detected during laparoscopy, and 3 of the 28 patients had a false-positive (n = 1) or a misleading positive (n = 2) lavage result. Therefore, lavage was beneficial in only 6/449 patients (1.3%); in these patients, the lavage result changed the assessment of tumor stage and adequately predicted irresectable disease. Univariate analysis showed a significant survival difference between patients in whom lavage detected tumor cells and those in whom it did not, but multivariate analysis revealed that these survival differences were caused by metastatic or ingrowing disease. CONCLUSION Cytology of peritoneal lavage with conventional staining should no longer be performed during laparoscopic staging of GI malignancies because it provides an additional benefit in only 1.3% of patients and has limited prognostic value for survival in this group of patients.

Alterations of the p53 tumor-suppressor gene and K-ras oncogene in perihilar cholangiocarcinomas from a high-incidence area.

We observed a clustering of cholangiocarcinoma in a part of West Virginia. We analyzed the frequency and type of alterations in the p53 tumor‐suppressor gene and the K‐ras oncogene to determine whether cholangiocarcinomas from this high‐incidence area differ from other cholangiocarcinomas at the molecular level. We studied 12 carcinomas of patients from the high‐incidence area (West Virginia group), and 15 carcinomas of patients from nearby states (non‐West Virginia group). Over‐expression of the p53 gene product, accompanying most mutations in the p53 gene, was determined by immunohistochemistry. p53 sequence analysis of exons 5, 6, 7, and 8 of the p53‐immunohistochemical‐positive carcinomas was also performed. K‐ras codon 12 mutations were detected by the polymerase chain reaction and allele‐specific oligonucleotide hybridization. Significantly more cholangiocarcinomas from the West Virginia group were p53‐immunohistochemical‐positive than from the non‐West Virginia group (67% vs. 20%; p < 0.05). p53 mutations did not differ in the 2 groups in respect to site or specific type. No differences were found between the 2 groups regarding K‐ras mutations (17% vs. 27%). Although the higher frequency of p53‐immunohistochemical positivity in the West Virginia group may reflect a different etiology of these cholangiocarcinomas, explaining the high incidence in this area, results of p53 sequence analysis were not different in the West Virginia group. The high incidence may be explained by difference in carcinogenic dose or a different etiology not reflected in p53 or K‐ras alterations. Int. J. Cancer 78:695–698, 1998.

Comparison of the novel quantitative ARMS assay and an enriched PCR–ASO assay for K-ras mutations with conventional cytology on endobiliary brush cytology from 312 consecutive extrahepatic biliary stenoses

Background: Extrahepatic biliary stenosis (EBS) has malignant and benign causes. Patients with EBS are at risk of having or developing malignancy. Accurate diagnostic tests for early detection and surveillance are needed. The sensitivity of biliary cytology for malignancy is low. K-ras mutation analysis on brush cytology is a valuable adjunct, but specificity is low. A quantitative test for K-ras mutations has been developed: the amplification refractory mutation system (ARMS). Aim: To assess the test characteristics and additional value of ARMS in diagnosing the cause of EBS. Methods: Brush samples from endoscopic retrograde cholangiopancreatography were collected from 312 patients with EBS. K-ras mutation analysis was performed using ARMS—allele specific amplification was coupled with real time fluorescent detection of PCR products. Results were compared with conventional cytology and K-ras mutation analysis using allele specific oligonucleotide (ASO) hybridisation, and evaluated in view of the final diagnosis. Results: The test characteristics of ARMS and ASO largely agreed. Sensitivity for detecting malignancy was 49% and 42%, specificity 93% and 88%, and positive predictive value (PPV) 96% and 91%, respectively. The sensitivity of ARMS and cytology combined was 71%, and PPV was 93%. The specificity of ARMS could be increased to 100% by setting limits for the false positives, but reduced sensitivity from 49% to 43%. Conclusions: ARMS can be considered supplementary to conventional cytology, and comparable to ASO in diagnosing malignant EBS. A specificity of 100% can be achieved with ARMS, which should be considered in the surveillance of patients at risk for pancreatic cancer.

Diagnostic p53 Immunostaining of Endobiliary brush cytology : Preoperative cytology compared with the surgical specimen

Endobiliary brush cytology is important in the distinction of malignant and benign causes of extrahepatic bile duct obstruction. The additional diagnostic value of p53 immunostaining on these cytology specimens was assessed.