Hereditary nonpolyposis colorectal cancer (HNPCC), also known as Lynch syndrome, is a common autosomal dominant syndrome characterized by early age at onset, neoplastic lesions, and microsatellite ...instability (MSI). Because cancers with MSI account for approximately 15% of all colorectal cancers and because of the need for a better understanding of the clinical and histologic manifestations of HNPCC, the National Cancer Institute hosted an international workshop on HNPCC in 1996, which led to the development of the Bethesda Guidelines for the identification of individuals with HNPCC who should be tested for MSI. To consider revision and improvement of the Bethesda Guidelines, another HNPCC workshop was held at the National Cancer Institute in Bethesda, MD, in 2002. In this commentary, we summarize the Workshop presentations on HNPCC and MSI testing; present the issues relating to the performance, sensitivity, and specificity of the Bethesda Guidelines; outline the revised Bethesda Guidelines for identifying individuals at risk for HNPCC; and recommend criteria for MSI testing.
Chronic infection and inflammation are risk factors for the development of cholangiocarcinoma, a highly malignant, generally fatal adenocarcinoma originating from biliary epithelia. However, the link ...between inflammation and carcinogenesis in these disorders is obscure. Because nitric oxide (NO) is generated in inflamed tissues by inducible nitric oxide synthase (iNOS) and because DNA repair proteins are potentially susceptible to NO-mediated nitrosylation, we formulated the hypothesis that inflammatory cytokines induce iNOS and sufficient NO to inhibit DNA repair enzymes leading to the development and progression of cholangiocarcinoma. iNOS and nitrotyrosine were demonstrated in 18/18 cholangiocarcinoma specimens. Furthermore, iNOS and NO generation could be induced in vitro by inflammatory cytokines (mixture of interleukin-1beta, IFN-gamma, and tumor necrosis factor alpha) in three human cholangiocarcinoma cell lines. NO-dependent DNA damage as assessed by the comet assay was demonstrated during exposure of the three cholangiocarcinoma cell lines to cytokines. Moreover, global DNA repair activity was inhibited by 70% by a NO-dependent process after exposure of cells to cytokines. Our data indicate that activation of iNOS and excess production of NO in response to inflammatory cytokines cause DNA damage and inhibit DNA repair proteins. NO inactivation of DNA repair enzymes may provide a link between inflammation and the initiation, promotion, and/or progression of cholangiocarcinoma.
BACKGROUND The incidence of oesophageal adenocarcinoma has increased greatly. Barrett's oesophagus is a known risk factor. AIMS To identify changes in the incidence, prevalence, and outcome of ...Barrett's oesophagus in a defined population. SUBJECTS Residents of Olmsted County, Minnesota, with clinically diagnosed Barrett's oesophagus, or oesophageal or oesophagogastric junction adenocarcinoma. METHODS Cases were identified using the Rochester Epidemiology Project medical records linkage system. Records were reviewed with follow up to 1 January 1998. RESULTS The incidence of clinically diagnosed Barrett's oesophagus (>3 cm) increased 28-fold from 0.37/100 000 person years in 1965–69 to 10.5/100 000 in 1995–97. Of note, gastroscopic examinations increased 22-fold in this same time period. The prevalence of diagnosed Barrett's oesophagus increased from 22.6 (95% confidence interval (CI) 11.7–33.6) per 100 000 in 1987 to 82.6/100 000 in 1998. The prevalence of short segment Barrett's oesophagus (<3 cm) in 1998 was 33.4/ 100 000. Patients with Barrett's oesophagus had shorter than expected survival but only one patient with Barrett's oesophagus died from adenocarcinoma. Only four of 64 adenocarcinomas occurred in patients with previously known Barrett's oesophagus. CONCLUSIONS The incidence and prevalence of clinically diagnosed Barrett's oesophagus have increased in parallel with the increased use of endoscopy. We infer that the true population prevalence of Barrett's oesophagus has not changed greatly, although the incidence of oesophageal adenocarcinoma increased 10-fold. Many adenocarcinomas occurred in patients without a previous diagnosis of Barrett's oesophagus, suggesting that many people with this condition remain undiagnosed in the community.
BACKGROUND: Microsatellite instability (MSI) and allelic imbalance involving chromosome arms 5q, 8p, 17p, and 18q are genetic alterations commonly found in colorectal cancer. We investigated whether ...the presence or absence of these genetic alterations would allow stratification of patients with Astler-Coller stage B2 or C colorectal cancer into favorable and unfavorable prognostic groups. METHODS: Tumors from 508 patients were evaluated for MSI and allelic imbalance by use of 11 microsatellite markers located on chromosome arms 5q, 8p, 15q, 17p, and 18q. Genetic alterations involving each of these markers were examined for associations with survival and disease recurrence. All P values are two-sided. RESULTS: In univariate analyses, high MSI (MSI-H), i.e., MSI at 30% or more of the loci examined, was associated with improved survival (P = .02) and time to recurrence (P = .01). The group of patients whose tumors exhibited allelic imbalance at chromosome 8p had decreased survival (P = .02) and time to recurrence (P = .004). No statistically significant associations with survival or time to recurrence were observed for markers on chromosome arms 5q, 15q, 17p, or 18q. In multivariate analyses, MSI-H was an independent predictor of improved survival (hazard ratio HR = 0.51; 95% confidence interval CI = 0.31-0.82; P = .006) and time to recurrence (HR = 0.42; 95% CI = 0.24-0.74; P = .003), and 8p allelic imbalance was an independent predictor of decreased survival (HR = 1.89; 95% CI = 1.25-2.83; P = .002) and time to recurrence (HR = 2.07; 95% CI = 1.32-3.25; P = .002). CONCLUSIONS: Patients whose tumors exhibited MSI-H had a favorable prognosis, whereas those with 8p allelic imbalance had a poor prognosis; both alterations served as independent prognostic factors. To our knowledge, this is the first report of an association between 8p allelic imbalance and survival in patients with colorectal cancer.
Background: Interstitial cells of Cajal (ICC) are required for normal intestinal motility. ICC are found throughout the human colon and are decreased in the sigmoid colon of patients with slow ...transit constipation. Aims: The aims of this study were to determine the normal distribution of ICC within the human colon and to determine if ICC are decreased throughout the colon in slow transit constipation. Patients: The caecum, ascending, transverse, and sigmoid colons from six patients with slow transit constipation and colonic tissue from patients with resected colon cancer were used for this study. Methods: ICC cells were identified with a polyclonal antibody to c-Kit, serial 0.5 μm sections were obtained by confocal microscopy, and three dimensional software was employed to reconstruct the entire thickness of the colonic muscularis propria and submucosa. Results: ICC were located within both the longitudinal and circular muscle layers. Two networks of ICC were identified, one in the myenteric plexus region and another, less defined network, in the submucosal border. Caecum, ascending colon, transverse colon, and sigmoid colon displayed similar ICC volumes. ICC volume was significantly lower in the slow transit constipation patients across all colonic regions. Conclusions: The data suggest that ICC distribution is relatively uniform throughout the human colon and that decreased ICC volume is pan-colonic in idiopathic slow transit constipation.
Frequent BRAF mutations were reported recently in a variety of human malignancies, including colorectal cancer. In this study, we screened 293 colorectal cancers for mutations in exons 11 and 15, two ...regions containing hotspots for BRAF mutation. Of the 293 cancers, 170 had normal mismatch repair, and 123 had defective mismatch repair (originating from both somatic as well as germ-line mutations in several of the mismatch repair genes). A total of 63 exonic mutations (22%) were detected, 60 of which were V599E, and one each of D593G, G468E, and D586A. Of the tumors with defective mismatch repair, 34% (42 of 123) had a mutation in BRAF, whereas only 12% (21 of 170) of tumors with proficient mismatch repair demonstrated a mutation (P < 0.0001). Interestingly, BRAF mutations were found most often in cases with an hMHL1 abnormality (35 of 60) and rarely in cases with an hMSH2 abnormality (1 of 39; P < 0.0001). More interestingly, of the 31 hMLH1 cases with a BRAF mutation, 30 occurred in tumors known to have hypermethylation of hMLH1 promoter. Only 1 of the 15 cases with a germ-line mutation in hMLH1 had a mutation in BRAF. In this series, BRAF mutations occurred rarely in tumors with defective mismatch repair attributable to the presence of germ-line mutation in either hMLH1 or hMSH2. Furthermore, BRAF mutations were strongly associated with the epigenetic alteration of hMLH1. Overall, these data suggest that BRAF mutations are not a consequence of defective mismatch repair per se.
Background & Aims: Cholestatic liver injury results from the intrahepatic accumulation of toxic bile salts. Toxic bile salt–induced hepatocyte apoptosis in vitro is Fas dependent. The aim of this ...study was to ascertain if hepatocyte apoptosis in vivo during cholestasis is Fas dependent.
Methods: Studies were performed in bile duct–ligated (BDL) Fas-deficient lpr (lymphoproliferation) and wild-type mice.
Results: Hepatocyte apoptosis was the predominant mechanism of cell death as determined by terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling and trypan blue assays to quantitate apoptosis and necrosis. The mechanisms of hepatocyte apoptosis were dependent on the presence or absence of the Fas receptor and the duration of BDL. After BDL of 3 days' duration, increased hepatocyte apoptosis occurred only in wild-type but not lpr mice, indicating the apoptosis was Fas dependent. In contrast, after BDL of ≥7 days, hepatocyte apoptosis also occurred in lpr animals consistent with a Fas-independent mechanism of apoptosis. Hepatocyte apoptosis in BDL lpr mice was associated with an increase in Bax expression and Bax association with mitochondria.
Conclusions: During extrahepatic cholestasis, hepatocyte apoptosis is mediated by Fas. However, in the absence of the Fas receptor, additional mechanisms of hepatocyte apoptosis occur. Inhibition of multiple apoptotic pathways is necessary to attenuate chronic cholestatic liver injury.
GASTROENTEROLOGY 1999;117:669-677
Recent studies have demonstrated the presence of microsatellite instability (MSI) in tumors from patients with hereditary nonpolyposis colon cancer and in a subset of patients with sporadic ...colorectal cancer (CRC). In sporadic CRC, three tumor phenotypes have been defined: microsatellite stable (MSS), low-frequency MSI, and high-frequency MSI (MSI-H). Although defective mismatch repair, consisting primarily of alterations in hMSH2 and hMLH1, is believed to be responsible for the MSI phenotype in the majority of patients with hereditary nonpolyposis colon cancer, the genetic defect responsible for this phenotype in sporadic CRC has yet to be clearly delineated. Somatic or germ-line alterations in these two genes have been identified in only a minority of these cases. Analysis of the protein expression patterns of hMSH2 and hMLH1 in unselected CRC, however, suggests that alterations in hMLH1 may account for a majority of the MSI-H cases. In an effort to explore the underlying molecular basis for these findings, we have examined the methylation status of the presumptive hMLHI promoter region in 31 tumors that vary in regard to their MSI status (MSI-H or MSS), their hMLH1 protein expression (MLH- or MLH+), and their gene mutation (Mut+ or Mut-) status. Hypermethylation of the hMLH1 promoter occurred in all 13 MSI-H/ MLH- tumors that did not have a detectable mutation within the hMLH1 gene. Of those MSI-H tumors containing germ-line or somatic alterations in hMLH1 (n = 7, including 3 frameshift, 1 nonsense, 2 missense mutations, and 1 tumor containing multiple mutations: missense, splice-site alteration, and a frameshift), four had a normal methylation pattern, whereas three others demonstrated hypermethylation of the hMLH1 promoter region. Two of these cases had a missense alteration, the other a frameshift alteration. The single MSI-H/Mut+ tumor that had normal hMLH1 and hMSH2 expression, as well as 9 of the 10 MSS cases, lacked methylation of the hMLH1 promoter. Hypermethylation of the hMSH2 promoter was not observed for any of the cases. These results suggest that hypermethylation of the hMLH1 promoter may be the principal mechanism of gene inactivation in sporadic CRC characterized by widespread MSI.
Background/Aims
: The pathogenesis of alcoholic hepatitis (AH) remains poorly understood. Although apoptosis is now recognized as a mechanism of liver injury, the extent and mechanisms of apoptosis ...in human AH remain unknown. Thus, our aims were to quantify hepatocyte apoptosis in patients with AH, correlate it with disease severity, and identify the mechanisms of apoptosis induction.
Methods
: Hepatocyte apoptosis was assessed in 26 patients with AH and 27 controls without liver disease using the TUNEL assay and immunohistochemistry for activated caspase 3. Liver specimens were also graded for disease severity. The expression of the death receptors, Fas and tumor necrosis factor-
α receptor 1 (TNF-R1), was assessed by immunohistochemistry.
Results
: In contrast to normal livers, TUNEL- and caspase 3-positive hepatocytes were readily observed in the livers of patients with AH. In the AH group, hepatocyte apoptosis was significantly higher in patients with a serum bilirubin of
>3 mg/dl. Apoptosis was also greater in grade 4 steatohepatitis. The Fas receptor was strongly expressed in hepatocytes in AH, but not in normal livers; the TNF-R1 expression was comparable in both groups.
Conclusions
: The present results demonstrate that hepatocyte apoptosis is significantly increased in human AH and justify therapeutic strategies aimed at inhibiting apoptosis in this disease.
Activation of Wnt signaling through beta-catenin mutations contributes to the development of hepatocellular carcinoma (HCC) and hepatoblastoma (HB). To explore the contribution of additional Wnt ...pathway molecules to hepatocarcinogenesis, we examined beta-catenin, AXIN1 and AXIN2 mutations in 73 HCCs and 27 HBs. beta-catenin mutations were detected in 19.2% (14 out of 73) HCCs and 70.4% (19 out of 27) HBs. beta-catenin mutations in HCCs were primarily point mutations, whereas more than half of the HBs had deletions. AXIN1 mutations occurred in seven (9.6%) HCCs and two (7.4%) HBs. The AXIN1 mutations included seven missense mutations, a 1 bp deletion, and a 12 bp insertion. The predominance of missense mutations found in the AXIN1 gene is different from the small deletions or nonsense mutations described previously. Loss of heterozygosity at the AXIN1 locus was present in four of five informative HCCs with AXIN1 mutations, suggesting a tumor suppressor function of this gene. AXIN2 mutations were found in two (2.7%) HCCs but not in HBs. Two HCCs had both AXIN1 and beta-catenin mutations, and one HCC had both AXIN2 and beta-catenin mutations. About half the HCCs with AXIN1 or AXIN2 mutations showed beta-catenin accumulation in the nucleus, cytoplasm or membrane. Overall, these data indicate that besides the approximately 20% of HCCs and 80% of HBs with beta-catenin mutations contributing to hepatocarcinogenesis, AXIN1 and AXIN2 mutations appear to be important in an additional 10% of HCCs and HBs.
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