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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.

Background: Juvenile polyps are distinct hamartomatous malformations of the gastrointestinal tract that may occur in the heritable juvenile polyposis syndrome (JPS) or sporadically. Histologically, juvenile polyps are characterized by a marked increase of the stromal cell compartment, but an epithelial phenotype has also been reported. JPS has an increased risk of colorectal cancer but sporadic juvenile polyps do not. In 50% to 60% of patients with JPS, a germline mutation of the transforming growth factor-β/bone morphogenetic protein (BMP) pathway genes SMAD4 or BMPR1A is found. This study compares the histologic phenotype of juvenile polyps with a SMAD4 or BMPR1A germline mutation and sporadic juvenile polyps., Methods: Hematoxylin and Eosin-stained slides of 65 JPS polyps and 25 sporadic juvenile polyps were reviewed for histologic features and dysplasia. Systematic random crypt and stroma counts were obtained by count stereology, and a crypt-stroma ratio was determined. All polyps were subsequently categorized as type A (crypt-stroma ratio <1.00) or type B (crypt-stroma ratio ≥1.00), the latter referring to the epithelial phenotype. Cell cycle activity was assessed using immunohistochemistry ofthe proliferation marker Ki67, and mutation analysis was carried out for KRAS and APC to determine the involvement of the adenoma-carcinoma sequence., Results: Juvenile polyps with a SMAD4 germline mutation were predominantly type B, whereas type A was more common among juvenile polyps with a BMPR1A germline mutation. However, this distinction could not be ascribed to differences in cell cycle activity. Dysplasia was equally common in JPS polyps with either a SMAD4 or BMPR1A germline mutation, in which the involvement of the adenoma-carcinoma sequence does not seem to play a distinct role., Conclusion: Juvenile polyps in the setting of JPS exhibit distinct phenotypes correlating with the underlying genetic defect.

Collision tumors are thought to arise from the accidental meeting of two independent tumors. Here we present five gastroesophageal junction tumors consisting of two collision tumors and three composite tumors (characterized by two divergent lineages originating from the same neoplastic clonal proliferation), as diagnosed on histology. In an attempt to prove this distinction at a genetic level, we performed TP53 sequence analysis and p53 immunohistochemistry. In addition, loss of heterozygosity (LOH) analysis using 10 microsatellite markers was carried out. An identical TP53 mutation and a similar pattern of retention and LOH were found in both neoplastic components of the presumed collision tumors, suggesting that both components are derived from a single precursor cell that undergoes divergent differentiation in the evolution of the tumor. In the composite group, 1 case had a genetic basis for the possible diagnosis of a collision tumor, with a TP53 mutation in the adenocarcinoma component only, and a different pattern of retention and loss of heterozygosity. These findings imply that it is not possible to recognize true collision tumors from immunohistologic appearance alone and suggest that the long-standing histologic criteria for the diagnosis of these neoplasms have no molecular basis.

Invasive pancreatic ductal adenocarcinoma is an almost uniformly fatal disease. Several distinct noninvasive precursor lesions can give rise to invasive adenocarcinoma of the pancreas, and the prevention, detection, and treatment of these noninvasive lesions offers the potential to cure early pancreatic cancers. Noninvasive precursors of invasive ductal adenocarcinoma of the pancreas include pancreatic intraepithelial neoplasias (PanINs), intraductal papillary mucinous neoplasms (IPMNs), and mucinous cystic neoplasms. Diagnostic criteria, including a distinct ovarian-type stroma, and a consistent nomenclature are well established for mucinous cystic neoplasms. By contrast, consistent nomenclatures and diagnostic criteria have been more difficult to establish for PanINs and IPMNs. Because both PanINs and IPMNs consist of intraductal neoplastic proliferations of columnar, mucin-containing cells with a variable degree of papilla formation, the distinction between these two classes of precursor lesions remains problematic. Thus, considerable ambiguities still exist in the classification of noninvasive neoplasms in the pancreatic ducts. A meeting of international experts on precursor lesions of pancreatic cancer was held at The Johns Hopkins Hospital from August 18 to 19, 2003. The purpose of this meeting was to define an international acceptable set of diagnostic criteria for PanINs and IPMNs and to address a number of ambiguities that exist in the previously reported classification systems for these neoplasms. We present a consensus classification of the precursor lesions in the pancreatic ducts, PanINs and IPMNs.

Rare pancreatic neoplasms have been reported that show both endocrine and exocrine differentiation in the neoplastic components. In addition, pancreatic endocrine tumors may contain small, cytologically bland ductules intimately admixed with the endocrine component. It was recently suggested that these ductules represent an intrinsic part of the tumor, ie, that the ductules are neoplastic, and the term "ductulo-insular tumors of the pancreas" was proposed. In the present study, the nature of the ductular component of 16 cases of ductule-containing pancreatic endocrine tumors was investigated at the molecular level. Molecular genetic changes often present in ductal pancreatic neoplasms were not found by immunohistochemistry for DPC4, p53, and ERBB2 and by sequence analysis of KRAS codon 12. An X-chromosome inactivation clonality assay of one such tumor from a female patient indicated that the neuroendocrine component was monoclonal, contrasting with the ductular component that was polyclonal. The lymph node and liver metastases from three patients only contained the neuroendocrine component, and no ductules were observed. Although certain morphologic features of ductule-containing endocrine tumors are reminiscent of the embryonic development of the human pancreas, none of the tumors expressed PDX-1, a transcription factor essential in pancreatic organ development. Based on our results, it is suggested that the ductular component occasionally found in pancreatic endocrine tumors is the result of entrapment of preexisting nonneoplastic ductules and that the tumors are otherwise not distinctive from conventional pancreatic endocrine tumors. Although the phenomenon is rare, it is important to recognize and to distinguish these tumors from true mixed ductal-endocrine neoplasms, which are generally more clinically aggressive. "Pancreatic endocrine tumors with entrapped ductules" would be the preferred nomenclature since it better reflects the nonneoplastic nature of the ductules.

Small cell carcinomas of the gallbladder are unusual neoplasms that have been characterized only recently. The authors describe the clinical, histopathologic, immunohistochemical, and molecular features of 12 small cell carcinomas of the gallbladder. The mean age at diagnosis was 69 years, and the male-to-female ratio was 5:7. The neoplasms had an average size of 3 cm, and 90% showed invasion of the muscularis propria and perimuscular connective tissue. Seventy-five percent of the carcinomas had metastasized or extended locally beyond the gallbladder at surgery. Survival was uniformly poor, with a mean survival of 10.7 months (range, 3-25 months). Half the small cell carcinomas were combined with other neoplasms. Four had foci of adenocarcinoma, one contained areas of squamous differentiation, and another had a component of carcinosarcoma. Immunohistochemical analysis showed focal reactivity for chromogranin (six of six cases), neuron-specific enolase (six of six cases), and Leu-7 (three of three cases). The molecular changes in small cell carcinomas were similar to those of adenocarcinomas occurring at this site, with a high frequency of p53 (75%) and p16INK4a (33%) abnormalities, and a low frequency of deleted in pancreatic carcinoma-4 inactivation (0%) and K-ras codon 12 mutations (17%). In contrast to pulmonary small cell carcinomas, p16INK4a function appears to be abrogated more frequently in these carcinomas.

Proliferative epithelial lesions in the smaller caliber pancreatic ducts and ductules have been the subject of numerous morphologic, clinical, and genetic studies; however, a standard nomenclature and diagnostic criteria for classifying these lesion have not been established. To evaluate the uniformity of existing systems for grading duct lesions in the pancreas, 35 microscopic slides with 35 representative duct lesions were sent to eight expert pathologists from the United States, Canada, and Europe. Kappa values for interobserver agreement could not be calculated initially because more than 70 different diagnostic terms were used by the eight pathologists. In several cases, the diagnoses rendered for a single duct lesion ranged from "hyperplasia," to "metaplasia," to "dysplasia," to "carcinoma in situ." This review therefore demonstrated the need for a standard nomenclature and classification system. Subsequently, during a working group meeting, the pathologists agreed to adopt a single standard system. The terminology pancreatic intraepithelial neoplasia (or PanIN) was selected, and diagnostic criteria for each grade of PanIN were established (http://pathology.jhu.edu/pancreas&#95;panin). This new system was then evaluated by having the eight pathologists rereview the original 35 cases. Only seven different diagnoses were rendered, and kappa values of 0.43, 0.14, and 0.42 were obtained for PanINs 1, 2, and 3 respectively. Cases assigned other diagnoses (e.g., squamous metaplasia) collectively had a kappa value of 0.41. These results show both the potential of the classification system, and also the difficulty of classifying these lesions even with a consistent nomenclature. However, even when there is lack of consensus, having a restricted set of descriptions and terms allows a better understanding of the reasons for disagreement. It is suggested that we adopt and apply this system uniformly, with continued study of its reliability and use, and possibly further refinement. The acceptance of a standard classification system will facilitate the study of pancreatic duct lesions, and will lead ultimately to a better understanding of their biologic importance.

The authors report seven patients with carcinoid tumors of the extrahepatic bile ducts (EHBDs). All patients were women, with an average age at diagnosis of 49.8 years (range, 37-67 yrs). The most common presenting symptom was painless jaundice with or without pruritus. Although one patient had peptic ulcer disease before the onset of obstructive jaundice, none had systemic endocrine manifestations. These neoplasms were most often located in the common bile duct. Grossly, the carcinoid tumors were usually nodular and poorly demarcated, and ranged from 1.1 to 2.7 cm in size. Only one of the neoplasms was polypoid. Microscopically, the tumors had a trabecular or nesting pattern with occasional tubule formation, and were composed of relatively small cells with granular chromatin. All of the neoplasms expressed chromogranin and two expressed synaptophysin. Three expressed serotonin and two of the three were also immunoreactive for pancreatic polypeptide or somatostatin. Two tumors were focally positive for gastrin and one of these two tumors was also positive for serotonin and pancreatic polypeptide. All seven carcinoid tumors showed no immunoreactivity for p53, and assays for p53 loss of heterozygosity analysis were negative in two, suggesting that p53 mutations do not play a role in the pathogenesis of EHBD carcinoids. A mutation in codon 12 of K-ras was found in one carcinoid tumor whereas two of two showed immunoreactivity for Dpc4 protein. In view of the small number of carcinoids studied, the importance of these findings in the pathogenesis of these tumors is unclear. Ultrastructural examination of three of the tumors revealed numerous membrane-bound, round neurosecretory granules. Clinically, these lesions had an indolent course. Even in the presence of lymph node metastases (noted in two patients), all of the patients remained disease free 2 to 11 years (average follow up, 6.6 yrs) after segmental resection or pancreaticoduodenectomy (Whipple's procedure). Because carcinoid tumors of the EHBD are of low malignant potential, they should be separated from the more common adenocarcinomas in this location.

A case of a pyloric gland type adenoma of the main pancreatic duct in a 69-year-old woman is reported. The tumor led to occlusion and cystic dilatation of the main duct in the pancreatic tail. The surgical resection specimen disclosed a polypoid, bilobed mass attached to the wall of the main pancreatic duct by a thin fibrous stalk. Light-microscopic examination revealed a well-demarcated nodule composed of closely packed tubular glands lined by columnar, mucin-secreting cells with abundant clear cytoplasm and basally oriented nuclei. Focal, mild cytologic atypia was seen. Pyloric metaplasia and focal papillary hyperplasia was present in the adjacent ductal epithelium. Periodic acid-Schiff reactions, with and without diastase predigestion, showed reactivity in the tubular glands, whereas alcian blue (pH 2.5) was negative. Immunohistochemical stains for chromogranin, serotonin, somatostatin, and gastrin failed to detect the respective antigens. Genetic analysis using polymerase chain reaction with mutant enrichment and allele specific oligonucleotide hybridization detected a single mutation at codon 12 of K-ras, which changed the wild-type glycine to arginine. This mutation is commonly found in invasive pancreatic ductal carcinomas. Although tumors with microscopic and immunohistochemical features consistent with pyloric gland adenoma have been described in the gallbladder, to our knowledge, this is the first reported case within the pancreatic ductal system. The finding of a K-ras, codon 12 mutation and the presence of focal dysplasia may denote neoplastic potential in association with this lesion.

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.

Mutations in the p53 tumor suppressor gene are frequently observed in primary lung adenocarcinomas, suggesting that these mutations are critical events in the malignant transformation of airway cells. These mutations are often associated with stabilization of the p53 gene product, resulting in the accumulation of p53 protein. In this study, 70 formalin-fixed, paraffin-embedded primary lung adenocarcinomas resected for potential cure were examined for p53 overexpression. These 70 lung adenocarcinomas were obtained from a series of patients with well-documented clinical histories, and all 70 carcinomas had been previously evaluated for point mutations in codon 12 of the K-ras oncogene. Overexpression of the p53 protein was detected using an antigen retrieval system (Target Unmasking Fluid) and the anti-p53 antibody CM-1. CM-1 is a polyclonal antibody directed against the wild-type p53 protein. Overexpression of the p53 protein was found in 23 (33%) of the 70 lung adenocarcinomas. In all 23 cases, overexpression was confined to neoplastic cells. Overexpression of the p53 protein correlated with cigarette smoking: 10 (56%) of the 18 adenocarcinomas from patients who were current smokers overexpressed p53 compared with 13 (33%) of the 40 adenocarcinomas from patients who had quit smoking and 0 (0%) of the 12 adenocarcinomas from patients who had never smoked (p = 0.002, trend test). Overexpression of the p53 protein was also related to the degree of histologic differentiation: 48% of the p53 negative carcinomas were well differentiated, whereas only 13% (p = 0.003) of the carcinomas in which p53 was overexpressed were well differentiated. Overexpression of the p53 protein did not correlate with point mutations in codon 12 of the K-ras oncogene, nor did it correlate with tumor stage or patient survival. These findings indicate that p53 protein is frequently overexpressed in primary lung adenocarcinomas. Furthermore, the association of tobacco smoking with this overexpression suggests that the p53 gene is a target of specific mutagens in tobacco smoke.