Lodewijk A.A. Brosens

Risk of colorectal cancer in juvenile polyposis

Background: Juvenile polyposis (JP) is an autosomal-dominant syndrome characterised by the development of hamartomatous gastrointestinal polyps and is associated with colorectal cancer. However, the relative and absolute risk of colorectal malignancy in these patients is not known. Methods: The incidence rates of colorectal cancer in patients with JP were compared with that of the general population through person-year analysis with adjustment for demographics. Results: In patients with JP, the RR (95% CI) of colorectal cancer was 34.0 (14.4 to 65.7). Similar risks were noted in both males (30.0, 9.6 to 68.6) and females (43.7, 8.8 to 125). The cumulative life-time risk for colorectal cancer was 38.7%. The mean (SD) age of diagnosis of colorectal cancer was 43.9 (10.4) years. Other gastrointestinal malignancies were not noted in this cohort. Conclusion: Patients with JP have a markedly increased RR and absolute risk for colorectal cancer and require vigilant colorectal surveillance starting at young age. A low threshold for recommending surgery with consideration for removal of the entire colorectum seems warranted.

A Revised Classification System and Recommendations From the Baltimore Consensus Meeting for Neoplastic Precursor Lesions in the Pancreas.

International experts met to discuss recent advances and to revise the 2004 recommendations for assessing and reporting precursor lesions to invasive carcinomas of the pancreas, including pancreatic intraepithelial neoplasia (PanIN), intraductal papillary mucinous neoplasm (IPMN), mucinous cystic neoplasm, and other lesions. Consensus recommendations include the following: (1) To improve concordance and to align with practical consequences, a 2-tiered system (low vs. high grade) is proposed for all precursor lesions, with the provision that the current PanIN-2 and neoplasms with intermediate-grade dysplasia now be categorized as low grade. Thus, “high-grade dysplasia” is to be reserved for only the uppermost end of the spectrum (“carcinoma in situ”–type lesions). (2) Current data indicate that PanIN of any grade at a margin of a resected pancreas with invasive carcinoma does not have prognostic implications; the clinical significance of dysplasia at a margin in a resected pancreas with IPMN lacking invasive carcinoma remains to be determined. (3) Intraductal lesions 0.5 to 1 cm can be either large PanINs or small IPMNs. The term “incipient IPMN” should be reserved for lesions in this size with intestinal or oncocytic papillae or GNAS mutations. (4) Measurement of the distance between an IPMN and invasive carcinoma and sampling of intervening tissue are recommended to assess concomitant versus associated status. Conceptually, concomitant invasive carcinoma (in contrast with the “associated” group) ought to be genetically distinct from an IPMN elsewhere in the gland. (5) “Intraductal spread of invasive carcinoma” (aka, “colonization”) is recommended to describe lesions of invasive carcinoma invading back into and extending along the ductal system, which may morphologically mimic high-grade PanIN or even IPMN. (6) “Simple mucinous cyst” is recommended to describe cysts >1 cm having gastric-type flat mucinous lining at most minimal atypia without ovarian-type stroma to distinguish them from IPMN. (7) Human lesions resembling the acinar to ductal metaplasia and atypical flat lesions of genetically engineered mouse models exist and may reflect an alternate pathway of carcinogenesis; however, their biological significance requires further study. These revised recommendations are expected to improve our management and understanding of precursor lesions in the pancreas.

Large genomic deletions of SMAD4, BMPR1A and PTEN in juvenile polyposis

Background/aims: Juvenile polyposis syndrome (JPS) is a rare autosomal dominant disorder characterised by multiple gastrointestinal juvenile polyps and an increased risk of colorectal cancer. This syndrome is caused by germline mutation of either SMAD4 or BMPR1A, and possibly ENG. PTEN, originally linked to Cowden syndrome and Bannayan–Riley–Ruvalcaba syndrome, has also been associated with JPS. By direct sequencing, germline mutations are found in only 30–40% of patients with a JPS phenotype. Therefore, alternative ways of inactivation of the known JPS genes, or additional genes predisposing to JPS may be involved. In this study, a comprehensive genetic analysis of SMAD4, BMPR1A, PTEN and ENG is performed through direct sequencing and multiplex ligation-dependent probe amplification (MLPA) in JPS patients. Methods: Archival material of 29 patients with JPS from 27 families was collected. Direct sequencing and MLPA analysis were performed to search for germline defects in SMAD4, BMPR1A, PTEN and ENG. Results: A germline defect in SMAD4, BMPR1A or PTEN was found in 13 of 27 (48.1%) unrelated JPS patients. Nine mutations (33.3%) were detected by direct sequencing, including six (22.2%) SMAD4 mutations and three (11.1%) BMPR1A mutations. MLPA identified four additional patients (14.8%) with germline hemizygous large genomic deletions, including one deletion of SMAD4, one deletion of exons 10 and 11 of BMPR1A, and two unrelated patients with deletion of both BMPR1A and PTEN. No ENG gene mutations were found. Conclusion: Large genomic deletions of SMAD4, BMPR1A and PTEN are a common cause of JPS. Using direct sequencing and MLPA, a germline defect was detected in 48.1% of JPS patients. MLPA identified 14.8% (4/27) of these mutations. Since a substantial percentage of JPS patients carry a germline deletion and MLPA is a reliable and user-friendly technique, it is concluded that MLPA is a valuable adjunct in JPS diagnosis.

Gastrointestinal Polyposis Syndromes

Colorectal cancer is one of the leading causes of cancer-related death in the Western society, and the incidence is rising. Rare hereditary gastrointestinal polyposis syndromes that predispose to colorectal cancer have provided a model for the investigation of cancer initiation and progression in the general population. Many insights in the molecular genetic basis of cancer have emerged from the study of these syndromes. This review discusses the genetics and clinical manifestations of the three most common syndromes with gastrointestinal polyposis and an increased risk of colorectal cancer: familial adenomatous polyposis (FAP), juvenile polyposis (JP) and Peutz-Jeghers syndrome (PJS).

SMAD4 Immunohistochemistry Reflects Genetic Status in Juvenile Polyposis Syndrome

Purpose: Juvenile polyposis syndrome (JPS) can be caused by a germline defect of the SMAD4 gene. Somatic inactivation of SMAD4 occurs in pancreatic and colorectal cancers and is reflected by loss of SMAD4 immunohistochemistry. Here, SMAD4 immunohistochemistry as a marker of SMAD4 gene status and the role of SMAD4 in the adenoma-carcinoma sequence in neoplastic progression in JPS are studied. Experimental Design: Twenty polyps with a SMAD4 germline defect and 38 control polyps were studied by SMAD4 immunohistochemistry. Inactivation of the SMAD4 wild-type allele was studied in dysplastic epithelium and in areas with aberrant SMAD4 expression. APC, β-catenin, p53, and K-ras were studied to evaluate the adenoma-carcinoma sequence. Results: Nine of 20 polyps with a SMAD4 germline defect showed loss of epithelial SMAD4 expression. Loss of heterozygosity of SMAD4 was found in five polyps and a somatic stop codon mutation was found in two polyps without loss of heterozygosity. Remarkably, somatic inactivation of epithelial SMAD4 did not always coincide with dysplasia and aberrant p53 staining was found in four of six dysplastic polyps with normal SMAD4 staining. One K-ras mutation was found in nine juvenile polyps with dysplasia. No evidence for Wnt activation was found. Conclusions: SMAD4 immunohistochemistry mirrors genetic status and provides a specific adjunct in the molecular diagnosis of JPS. However, epithelial SMAD4 inactivation is not required for polyp formation and is not obligatory for neoplastic progression in JPS. Instead, different routes to neoplasia in JPS caused by germline SMAD4 mutation seem to be operative, including somatic loss of SMAD4 and p53 inactivation without somatic loss of SMAD4. Clin Cancer Res; 16(16); 4126–34. ©2010 AACR.

Analysis of LKB1 mutations and other molecular alterations in pancreatic acinar cell carcinoma

Acinar cell carcinoma is a rare non-ductal neoplasm of the pancreas with poorly defined molecular genetic features. Recently, biallelic inactivation of LKB1 was described in an acinar cell carcinoma of a Peutz-Jeghers patient carrying a heterozygous germline LKB1 mutation, and inhibition of mTOR signaling resulted in partial remission of the tumor. To explore the potential of mTOR inhibitors in sporadic acinar cell carcinoma, the LKB1 gene was investigated in five sporadic acinar cell carcinomas by sequence analysis, methylation analysis and mRNA expression. In addition, microsatellite instability and methylation of a number of tumor suppressor genes were investigated and KRAS, TP53, CDKN1A, SMAD4 and CTNNB1 were studied by mutation analysis and immunohistochemistry. No mutations, deletions or promoter hypermethylation of LKB1 were found in any of the sporadic acinar cell carcinomas, and mRNA expression of LKB1 was not altered. Amplifications at chromosome 20q and 19p were found in 100 and 80% of the cases, respectively. In addition, hypermethylation of one or more tumor suppressor genes was found in 80% of cases. One case harbored a TP53 mutation, and expression of SMAD4 and CTNNB1 was altered in one case each. No KRAS mutations or microsatellite instability were found. To conclude, no evidence for a role for LKB1 in tumorigenesis of sporadic pancreatic acinar cell carcinoma was found. However, copy number variations and hypermethylation were found in a majority of cases. Molecular pathways involved in acinar cell carcinoma-tumorigenesis differ from those involved in ductal pancreatic neoplasms. Further studies are needed to increase our understanding of molecular pathogenesis of acinar cell carcinoma, which may eventually result in development of new therapeutic targets.

Focal nodular hyperplasia: hepatobiliary enhancement patterns on gadoxetic-acid contrast-enhanced MRI

ObjectivesTo assess the range of hepatobiliary enhancement patterns of focal nodular hyperplasia (FNH) after gadoxetic-acid injection, and to correlate these patterns to specific histological features.Materials and methodsFNH lesions, imaged with Gadoxetic-acid-enhanced MRI, with either typical imaging findings on T1, T2 and dynamic-enhanced sequences or histologically proven, were evaluated for hepatobiliary enhancement patterns and categorized as homogeneously hyperintense, inhomogeneously hyperintense, iso-intense, or hypo-intense-with-ring. Available histological specimens of FNHs (surgical resection or histological biopsy), were re-evaluated to correlate histological features with observed enhancement patterns.Results26 FNHs in 20 patients were included; histology was available in six lesions (four resections, two biopsies). The following distribution of enhancement patterns was observed: 10/26 homogeneously hyperintense, 4/26 inhomogeneously hyperintense, 5/26 iso-intense, 6/26 hypointense-with-ring, and 1/26 hypointense, but without enhancing ring. The following histological features associated with gadoxetic-acid uptake were identified: number and type of bile-ducts (pre-existent bile-ducts, proliferation, and metaplasia), extent of fibrosis, the presence of inflammation and extent of vascular proliferation.ConclusionFNH lesions can be categorized into different hepatobiliary enhancement patterns on Gadoxetic-acid-enhanced MRI, which appear to be associated with histological differences in number and type of bile-ducts, and varying the presence of fibrous tissue, inflammation, and vascularization.

Surgical and molecular pathology of pancreatic neoplasms.

BackgroundHistologic characteristics have proven to be very useful for classifying different types of tumors of the pancreas. As a result, the major tumor types in the pancreas have long been classified based on their microscopic appearance.Main bodyRecent advances in whole exome sequencing, gene expression profiling, and knowledge of tumorigenic pathways have deepened our understanding of the underlying biology of pancreatic neoplasia. These advances have not only confirmed the traditional histologic classification system, but also opened new doors to early diagnosis and targeted treatment.ConclusionThis review discusses the histopathology, genetic and epigenetic alterations and potential treatment targets of the five major malignant pancreatic tumors - pancreatic ductal adenocarcinoma, pancreatic neuroendocrine tumor, solid-pseudopapillary neoplasm, acinar cell carcinoma and pancreatoblastoma.

Increased expression of cytoplasmic HuR in familial adenomatous polyposis

Background: HuR is an mRNA stability factor that binds to the AU-rich element-containing 3’ untranslated region of the transcript. HuR overexpression is associated with increased tumor growth. Increased cytoplasmic HuR expression occurs in several cancer types, including colorectal cancer where it may contribute to the increased cyclooxygenase-2 (COX-2) expression observed during tumorigenesis. To investigate expression of HuR in the colorectal adenoma-carcinoma sequence, we examined expression of HuR in colorectal mucosa of patients with familial adenomatous polyposis (FAP) and sporadic colorectal cancer with correlation to COX-2 expression. Materials and methods: HuR and COX-2 protein expression were studied by immunohistochemistry of normal colon mucosa (N=20), adenomas (N=112), carcinomas (N=9) from patients with FAP, and 141 sporadic colorectal adenocarcinomas (Dukes B and C). Results: Cytoplasmic HuR staining was found in the epithelium of 10% of normal mucosa, 14.3% of adenomas and 88.9% of adenocarcinomas from FAP patients (p < 0.01) and in 68.8% of sporadic colorectal carcinomas. High epithelial COX-2 immunostaining was observed in 10% of normal, 8% of adenomas and all adenocarcinomas from FAP patients (p < 0.01) and in 69.5% of sporadic colorectal carcinomas. Positive cytoplasmic HuR immunostaining correlated with high COX-2 immunoreactivity in colon mucosa of FAP patients (p < 0.01) and in sporadic colorectal carcinomas. (p = 0.016)Conclusions: HuR is increasingly expressed in the cytoplasmic epithelial compartment in consecutive stages of the adenoma-carcinoma sequence in FAP. Also, COX-2 levels correlate with cytoplasmic expression of HuR in colonic epithelium of FAP patients and in sporadic colorectal cancer specimens. The role of cytoplasmic expression of HuR as a biomarker for progression of adenomas in FAP needs further study.

Genetic analyses of isolated high-grade pancreatic intraepithelial neoplasia (HG-PanIN) reveal paucity of alterations in TP53 and SMAD4

High‐grade pancreatic intraepithelial neoplasia (HG‐PanIN) is the major precursor of pancreatic ductal adenocarcinoma (PDAC) and is an ideal target for early detection. To characterize pure HG‐PanIN, we analysed 23 isolated HG‐PanIN lesions occurring in the absence of PDAC. Whole‐exome sequencing of five of these HG‐PanIN lesions revealed a median of 33 somatic mutations per lesion, with a total of 318 mutated genes. Targeted next‐generation sequencing of 17 HG‐PanIN lesions identified KRAS mutations in 94% of the lesions. CDKN2A alterations occurred in six HG‐PanIN lesions, and RNF43 alterations in five. Mutations in TP53, GNAS, ARID1A, PIK3CA, and TGFBR2 were limited to one or two HG‐PanINs. No non‐synonymous mutations in SMAD4 were detected. Immunohistochemistry for p53 and SMAD4 proteins in 18 HG‐PanINs confirmed the paucity of alterations in these genes, with aberrant p53 labelling noted only in three lesions, two of which were found to be wild type in sequencing analyses. Sixteen adjacent LG‐PanIN lesions from ten patients were also sequenced using targeted sequencing. LG‐PanIN harboured KRAS mutations in 94% of the lesions; mutations in CDKN2A, TP53, and SMAD4 were not identified. These results suggest that inactivation of TP53 and SMAD4 are late genetic alterations, predominantly occurring in invasive PDAC. Copyright