David N. Lewin

Lymphovascular Invasion in Colorectal Cancer: An Interobserver Variability Study

Background Lymphovascular invasion (LVI) in colorectal cancer (CRC) is considered a strong stage-independent prognostic factor and influences decisions regarding adjuvant chemotherapy in patients with stage II tumors. However, the degree of interobserver agreement among pathologists for LVI in CRC is largely unknown. This study was undertaken to examine such interobserver variability, and we hypothesized that the use of immunohistochemical markers for vascular and lymphatic channels could improve interobserver agreement. Design Fifty cases of American Joint Committee on Cancer stage II moderately differentiated CRC from 1990 to 2005 from the pathology archives were selected; mucinous, medullary, and other recognized special subtypes were excluded. Fifty hematoxylin and eosin (H&E) slides (1 from each case) were circulated to 6 gastrointestinal pathologists, who independently assessed small and large vessel invasion. No diagnostic guidelines were given to the participating pathologists; each was instructed to apply the criteria for LVI that he or she used in daily practice. Immunohistochemistry (IHC) for D2-40 and CD31 was performed on corresponding paraffin blocks. The IHC slides were randomized, recirculated, and rescored for LVI. Results were analyzed by kappa (κ) statistics, which correct for agreement by chance, and for percentage agreement. Results The average κ values were determined for the H&E slides (large and small vessel), CD31 (small vessel), and D2-40 (small vessel) (Fig. 1). Agreement was fair for H&E small vessel invasion [κ=0.28; 95% confidence interval (CI): 0.22-0.34]. The least agreement was seen in interpretation of H&E large vessel invasion (κ=0.18; 95%CI: 0.11-0.26). Agreement was not improved by use of immunohistochemical stains: CD31 (large vessel, κ=0.42, 95%CI: 0.20-0.63, small vessel, κ=0.26, 95%CI: 0.10-0.42) and D2-40 (κ=0.32, 95%CI: 0.21-0.42). Conclusions Interobserver variability in diagnosis of LVI was substantial on H&E slides and did not improve upon use of IHC. Agreement in evaluation of large vessel invasion was only slightly higher than would be seen by chance alone. This study highlights the need for criteria in evaluation of LVI, as this assessment may impact patient prognosis and thus change the course of clinical treatment.

Colonoscopic miss rates determined by direct comparison of colonoscopy with colon resection specimens

Colonoscopic miss rates determined by direct comparison of colonoscopy with colon resection specimens

Evaluation of gastric biopsies for neoplasia : Differences between Japanese and western pathologists

Cited variations in the evaluation of gastric endoscopic biopsies for neoplasms between pathologists in Japan and those in the United States and Europe (the West) may have stemmed from several causes. The five-tiered group classification of the Japanese Research Society for Gastric Cancer (JRSGC) for interpretation of biopsies is not used in the West. Some differences may also exist in the morphologic criteria to reach a diagnosis of dysplasia or carcinoma. The goals of this study were to test the Western and Japanese classifications of gastric dysplasia and adenocarcinoma and to assess the differences between four Japanese and seven Western pathologists. One hundred biopsies, 20 from each of the five categories of the JRSGC scheme as determined by one observer, were collected. The Japanese observers used the JRSGC system, expressed in Roman numerals, whereas Western pathologists used a five- or six-tiered scheme expressed in diagnostic terms. Pairwise agreement was evaluated using k statistics within both groups. Consensus diagnosis on each biopsy was accepted as the opinion of the majority. The sensitivity and specificity of each reviewer for a certain diagnosis were also assessed. The intragroup agreements were moderate for both the Japanese (mean k = 0.663) and the Westerners (mean k = 0.652). The pairwise agreements between Japanese and Western observers were low (mean k = 0.542). Overall, the sensitivity was low for all Japanese observers for the diagnosis of dysplasia (38.7% vs 92.5%), and the sensitivity for the diagnosis of adenocarcinoma was high in both groups but higher among the Japanese observers (93.9% and 85.2%, respectively). Overall, the Japanese-Western interobserver agreement was moderate. The JRSCG scheme did not translate into higher interobserver agreement among Japanese observers. The sensitivity for the diagnosis of gastric adenocarcinoma was high for both groups, but the specificity was low among the Japanese. The cause seemed to be centered around the diagnosis of dysplasia in the Western system, which was a lesion frequently interpreted as carcinoma in Japan because of the different definitions of carcinoma in each system. Such a discrepancy might be important because it may explain some of the differences in the prevalence and prognosis of early gastric cancer between Japan and the West. An international effort is needed to harmonize morphologic criteria and analyze whether therapeutic consequences may stem from such discrepancies.

Accurate discrimination of pancreatic ductal adenocarcinoma and chronic pancreatitis using multimarker expression data and samples obtained by minimally invasive fine needle aspiration

To augment cytological diagnosis of pancreatic ductal adenocarcinoma (PDAC) in tissue samples obtained by minimally invasive endoscopic ultrasound‐guided fine needle aspiration, we investigated whether a small set of molecular markers could accurately distinguish PDAC from chronic pancreatitis (CP). Expression levels of 29 genes were first determined by quantitative real‐time RT‐PCR in a training set of tissues in which the final diagnosis was PDAC (n = 20) or CP (n = 10). Using receiver operator characteristic curve analysis, we determined that the single gene with the highest diagnostic accuracy for discrimination of CP vs. PDAC in the training study was urokinase plasminogen activator receptor (UPAR; AUC value = 0.895, 95% CI = 0.728–0.976). In the set of test tissues (n = 14), the accuracy of UPAR decreased to 79%. However, we observed that the addition of 6 genes (EPCAM2, MAL2, CEA5, CEA6, MSLN and TRIM29; referred to as the 6‐gene classifier) to UPAR resulted in high accuracy in both training and testing sets. Excluding 3 samples (out of 44; 7%) for which results of the UPAR/6‐gene classifier were “undefined,” the accuracy of the UPAR/6‐gene classifier was 100% in training samples (n = 29), 92% in 12 test samples (p = 0.004 that results were randomly generated; p = 0.046 that the UPAR/6‐gene classifier was comparable to UPAR alone; χ2 test), 100% in 3 samples for which the initial cytological diagnosis was “suspicious” and 98% (40/41) overall. Our results provide evidence that molecular marker expression data can be used to augment cytological analysis.

Development of an unbiased method for the estimation of liver steatosis

Abstract:  Background:  Steatosis significantly contributes to an organs transplantability. Livers with >30% fat content have a 25% chance of developing primary non‐function (PNF). The current practice of evaluating a hematoxylin and eosin (H&E) stained donor biopsy by visual interpretation is subjective. We hypothesized that H&E staining of frozen sections fails to accurately estimate the degree of steatosis present within a given liver biopsy. To address this problem of evaluating steatosis in prospective donor organs, we developed a fast, user friendly computer methodology to objectively assess fat content based on the differential quantification of color pixels in Oil Red O (ORO) stained liver biopsies.

Telecytopathology for immediate evaluation of fine-needle aspiration specimens†

On‐site evaluation of fine‐needle aspiration (FNA) specimens by a pathologist is essential to obtain adequate samples and provide a preliminary diagnosis. Distance from the laboratory can make this difficult. The authors present their experience with on‐site evaluation using telecytopathology.

Liver Biopsy Results in Patients with Sickle Cell Disease on Chronic Transfusions: Poor Correlation With Ferritin Levels

Chronic transfusions are effective in preventing stroke and other complications of sickle cell disease. The aim of this study was to determine whether serum ferritin levels correlated with liver iron content in sickle cell patients on chronic transfusion.

Anti‐Endotoxin Monoclonal Antibodies are Protective against Hepatic Ischemia/Reperfusion Injury in Steatotic Mice

Steatotic mice are particularly susceptible to hepatic ischemia/reperfusion injury compared with their lean littermates. We have previously demonstrated that livers of mice having a spontaneous mutation in the leptin gene (ob/ob), resulting in global obesity and liver steatosis, are ATP depleted, are endotoxin sensitive, and do not survive (I/R) injury. We hypothesize that administration of an anti‐LPS monoclonal antibody (mAb) prior to initiation of I/R would be protective from that insult. Steatotic mice (ob/ob) were subjected to 15 min of ischemia via complete porta‐hepatis occlusion and varying lengths of reperfusion with or without pre‐treatment with an anti‐LPS mAb. There was 14–31% survival of isotype matched control mAb treated ob/ob mice after 15 min of ischemia and 24 h of reperfusion. In contrast, 75–83% of ob/ob mice pre‐treated with an anti‐LPS mAb prior to initiation of I/R survived both ischemia and 24 h of reperfusion. Furthermore, there was a decrease in ALT and circulating endotoxin levels when treated with an anti‐LPS mAb compared with control antibodies. Attenuation of the endotoxin load with anti‐LPS mAb, prior to initiation of I/R, was cytoprotective and improved survival. Consequently, these studies might offer a solution to the problems associated with using steatotic livers in clinical transplantation.

Are Ulcers a Marker for Invasive Carcinoma in Barrett's Esophagus? Data From a Diagnostic Variability Study With Clinical Follow-Up

OBJECTIVES: We correlated follow-up information from 138 patients with Barrett’s esophagus and varying degrees of dysplasia with the presence of ulcers. METHODS: A group of pathologist participants were asked to contribute patients’ initial biopsy slides showing Barrett’s esophagus (BE) without dysplasia and with epithelial changes indefinite for dysplasia, low grade dysplasia (LGD), high grade dysplasia (HGD), and adenocarcinoma. From the initial 250 cases used for a diagnostic reproducibility study, follow-up information was available for 138 patients. RESULTS: There were 44 cases submitted as BE, 22 as BE with epithelial changes indefinite for dysplasia, 26 as BE with LGD, 33 as BE with HGD, and 13 as BE with adenocarcinoma. Ulcers were present in 35/138 cases (25%), including 3/44 cases of BE without dysplasia (7%), 2/22 cases of BE with epithelial changes indefinite for dysplasia (9%), 0/26 cases of BE with LGD (0%), 10/33 cases of BE with HGD (30%), and 7/13 cases of BE with adenocarcinoma (54%). On follow-up, there were no invasive carcinomas detected among the BE without dysplasia group (median follow-up 38.5 months). Adenocarcinomas were detected in 4/22 cases (18%) submitted as BE with epithelial changes indefinite for dysplasia at 19, 55, 60, and 62 months and in 4/26 cases (15%) of BE with LGD at 9, 9, 11, and 60 months. None of these carcinomas occurred in cases in which an ulcer was present in the initial biopsy specimen. Among the 33 HGD cases, 20 (60%) were found to have adenocarcinoma on subsequent resection specimens. The presence of an ulcer with HGD increased the likelihood of finding carcinoma in the resection specimen, as 8/10 biopsies (80%) of HGD patients with ulcers had carcinoma, compared to 12/23 biopsies (52%) of HGD patients without ulcers. All of the cases interpreted as adenocarcinomas on biopsy were found either to have invasive carcinoma on esophageal resection or to have metastases that were demonstrated in unresectable patients. CONCLUSION: If an ulcer accompanies HGD in a biopsy specimen from a patient with BE, it is likely that invasive carcinoma is also present at that time. (Am J Gastroenterol 2002;97: 27–31.

Analysis of Protein Expression and Gene Mutation of c-kit in Colorectal Neuroendocrine Carcinomas

Primary neuroendocrine carcinomas of the colon are rare but highly aggressive malignancies. The recent observations that c-kit protooncogene, a tyrosine kinase, is overexpressed in a subset of small cell lung cancer and that selective kinase inhibitors block the in vitro growth of small cell lung cancer cell lines prompted us to investigate the expression and mutation status of the c-kit gene in colorectal neuroendocrine carcinomas. Sixty-six cases of primary colorectal neuroendocrine carcinoma were collected from 13 institutions, including 36 small cell carcinomas and 30 moderately differentiated neuroendocrine carcinomas. Immunohistochemical studies using a polyclonal antibody against c-kit protein (CD117) demonstrated a strong and diffuse cytoplasmic staining in 15 cases (23%), which were relatively equally distributed in the small cell and moderately differentiated subgroups. As controls, 25 conventional colorectal adenocarcinomas, 26 colorectal adenomas and 19 colorectal carcinoid tumors were all negative, whereas 15 gastrointestinal stromal tumors were all positive, for kit expression. In contrast to gastrointestinal stromal tumors, kit-overexpressing neuroendocrine carcinomas showed no mutations in the juxtamembrane domain (exon 11) of the c-kit gene as determined by mutational analysis. Kaplan-Meier analysis with the log-rank test revealed that the patients with kit-positive tumors did not differ significantly in survival from those with kit-negative tumors (P = 0.77). These results indicate that c-kit overexpression observed in a subset of colorectal neuroendocrine carcinomas may not be mediated via activating mutations, and does not appear to be an initiating event during tumorigenesis because of lack of c-kit expression in other types of colorectal epithelial neoplasms. More importantly, our observations may have potential therapeutic implications since specific tyrosine kinase inhibitors have shown promise in the management of patients with kit-expressing malignancies.