Abstract

Context.—Metastatic disease is the most critical determinant of resectability of pancreatic cancer and accounts for the poor outcome of patients with this disease. Thus, a better understanding of metastatic pancreatic cancer will afford new opportunities for therapeutic intervention.

Objective.—To summarize and discuss the current understanding of the clinical and molecular features of metastatic pancreatic cancer.

Data Sources.—Published literature on advanced stage pancreatic cancer, pancreatic cancer metastasis, and autopsy findings in patients with pancreatic cancer.

Conclusions.—In the clinical setting, it can be difficult to distinguish a metastatic pancreatic carcinoma from primary neoplasms in the liver, lung, or ovary. However, immunolabeling for DPC4 protein as part of a diagnostic panel is useful for making this distinction. Emerging data from a variety of investigators now indicate that overexpression of EphA2, loss of DPC4 and MKK4, and aberrant activation of the Hedgehog signaling pathway are associated with metastatic propensity of pancreatic cancers, providing novel therapeutic targets for the most lethal stage of this disease.

Pancreatic cancer is the fourth leading cause of cancer-related mortalities in the United States. In the year 2008 in the United States, an estimated 37 680 patients will be diagnosed with pancreatic cancer, and 34 290 patients will die of their disease.1 These unfortunate statistics reflect the advanced stage at which most patients with pancreatic cancer are diagnosed and the paucity of effective chemotherapeutic regimens for advanced disease.2–4 Thus, a better understanding of the factors that contribute to pancreatic cancer progression and metastasis is critical in combating this disease.

Contrary to early pancreatic carcinogenesis,5 the pathology and molecular features of advanced stage pancreatic cancers are relatively unexplored. As a result, little is known about the mechanisms responsible for uncontrolled local growth and metastatic spread, the very processes ultimately responsible for most pancreatic cancer– related deaths.6 This is underscored by the fact that unlike advanced stage colorectal cancers in which the primary cancer and/or liver metastases are commonly resected for clinical benefit and are thus available for medical research, the management of advanced stage pancreatic cancer often does not include surgical resection.7,8 

A better understanding of the clinical, genetic, and molecular features of metastatic pancreatic cancer will afford novel areas for therapeutic intervention. Toward this goal, we provide a review of the literature on the pathology and genetics of advanced stage pancreatic cancer including recent advances that may lead to novel treatments for this most lethal stage of the disease.

GROSS FEATURES

Distant metastases are a frequent occurrence in patients with pancreatic cancer.9–12 The most common site of distant metastasis found at autopsy is the liver, followed by the peritoneum, lung and pleura, bones, and adrenal glands (Figure 1).9–12 However, metastatic pancreatic cancer has been reported in virtually every organ site, including the brain, diaphragm, gallbladder, heart, small and large intestines, kidneys, ovaries, pericardium, seminal vesicles, skin, stomach, spleen, testis, thyroid gland, urinary bladder, and uterus.9,10,12 Calculations based on estimates of tumor doubling times indicate that occult liver metastases are often present at the time of pancreatectomy.13 Even small (<2 cm) pancreatic cancers are associated with metastases,10 suggesting metastatic propensity is rapidly acquired during pancreatic carcinogenesis and progression. Consistent with this notion, molecular analyses indicate micrometastases are frequently present within the peritoneal cavity and bone marrow of patients with potentially resectable pancreatic cancer.14–16 

Figure 1.

Frequency of metastatic involvement by pancreatic cancer to various organ sites. The liver is the most common site of metastasis, followed by the peritoneum and lung. Adrenal and bone metastases consistently account for ∼10% of metastatic disease from pancreatic cancer. LN indicates lymph node

Figure 1.

Frequency of metastatic involvement by pancreatic cancer to various organ sites. The liver is the most common site of metastasis, followed by the peritoneum and lung. Adrenal and bone metastases consistently account for ∼10% of metastatic disease from pancreatic cancer. LN indicates lymph node

Although the liver is the most common site of distant metastasis, up to one-third of patients do not have hepatic metastases at autopsy. In these patients, the lungs and abdominal lymph nodes are the most common sites of metastasis possibly due to hepatofugal portosystemic shunting (ie, flow directed away from the liver to the systemic circulation) induced by splenic vein obstruction or to the presence of aggressive features of the primary carcinoma.9–11 Mao et al10 found that carcinomas of the body and/ or tail of the pancreas are more frequently associated with hematogenous dissemination than are carcinomas of the head of the pancreas. Independent of the variability among patients with respect to the sites of metastatic spread, the extent of metastatic disease at autopsy is also widely variable, ranging from less than 10 documented metastases to more than 1000 metastases.9–11,17 

Hishinuma et al18 analyzed recurrence patterns for 25 autopsied patients who had undergone potentially curative resection of pancreatic cancer. Three-fourths of the patients had a local recurrence, half had hepatic metastasis, and one-third developed peritoneal dissemination. Overall, distant metastasis at any site was recognized in three-fourths of the patients. In our experience, 73% of patients who initially presented with stage I/II disease and underwent surgical resection of their infiltrating carcinomas had gross evidence of local recurrence, and two-thirds of these patients had metastatic disease.19 Although local recurrence frequently occurs following resection of pancreatic cancer,20,21 its direct relationship to patient survival is unclear.22,23 

MICROSCOPIC FEATURES

Kamisawa et al9 investigated the histologic features of pancreatic cancers in 130 patients with and without associated metastatic disease at autopsy and found that the primary cancer in patients with distant metastases tended to be more poorly differentiated. Moreover, we and others have found that advanced stage pancreatic cancers more commonly have high-grade (anaplastic) features than those reported for early stage disease.3,9,19 Of note, the finding of high-grade features at autopsy is not correlated with clinical stage or grade at initial diagnosis, indicating loss of tumor differentiation occurs with disease recurrence and/or progression.19 Winter et al24 found that loss of E-cadherin expression in pancreatic cancer is almost universally associated with anaplastic features (Figure 2), but this was not due to a methylation of the E-cadherin promoter with disease progression. In vitro models suggest that loss of differentiation in pancreatic cancer, that is epithelial-mesenchymal transition, is mediated by transforming growth factor β (TGF-β) signaling through the phosphoinositide 3-kinase/phosphatase and tensin homolog pathway.25–27 

Figure 2.

Example of recurrent high-grade pancreatic cancer identified at rapid autopsy. A, This patient underwent surgical resection of a moderately differentiated duct (tubular) adenocarcinoma and completed adjuvant chemoradiation. The carcinoma recurred 21 months postresection (hematoxylin-eosin, original magnification ×200). B, At autopsy, widespread undifferentiated anaplastic carcinoma was found in this same patient that accounted for more than 90% of the neoplastic cellularity. The cells show marked nuclear enlargement and pleomorphism with complete loss of cellular adhesion (hematoxylin-eosin, original magnification ×200). C, E-cadherin labeling of the carcinoma shown in B indicates the loss of E-cadherin expression in the undifferentiated region, in striking contrast to the focus of residual moderately differentiated carcinoma present in the same section (original magnification ×200)

Figure 2.

Example of recurrent high-grade pancreatic cancer identified at rapid autopsy. A, This patient underwent surgical resection of a moderately differentiated duct (tubular) adenocarcinoma and completed adjuvant chemoradiation. The carcinoma recurred 21 months postresection (hematoxylin-eosin, original magnification ×200). B, At autopsy, widespread undifferentiated anaplastic carcinoma was found in this same patient that accounted for more than 90% of the neoplastic cellularity. The cells show marked nuclear enlargement and pleomorphism with complete loss of cellular adhesion (hematoxylin-eosin, original magnification ×200). C, E-cadherin labeling of the carcinoma shown in B indicates the loss of E-cadherin expression in the undifferentiated region, in striking contrast to the focus of residual moderately differentiated carcinoma present in the same section (original magnification ×200)

Terayama et al28 investigated the pathology of liver metastases derived from a variety of primary sites and found that small pancreatic cancer metastases frequently show an intraparenchymal, expansive pattern of growth compared with other tumor types. However, irrespective of the primary carcinoma it was derived from, the proportion of metastases showing an expansive growth pattern increased in relation to the size of the metastatic deposit. A fibrous capsule around liver metastases, which is commonly observed in liver metastases from colorectal cancer,29 was not seen in any liver metastases derived from pancreatic cancers.

DIFFERENTIAL DIAGNOSIS

In the clinical setting, pancreatic cancer metastases can pose a diagnostic dilemma in a variety of organ sites. In the liver, small metastases need to be distinguished from benign epithelial lesions, including bile duct adenomas and bile duct hamartomas (Figure 3). Bile duct adenomas are small (<1 cm), subcapsular, circumscribed, noncystic solitary lesions composed of uniform cuboidal cells that form tubules.30,31 They are believed to be a reactive process to a focal injury.30 By contrast, bile duct hamartomas (von Meyenburg complexes) are usually multiple, are cystic, contain bile, and are lined by flattened uniform “biliary” type epithelial cells.32 In the event that routine histopathologic analysis is unable to distinguish between these benign entities and a metastasis from a pancreatic cancer, immunohistochemical labeling for a variety of markers may be of value. For example, Hornick et al33 found that abnormal p53 labeling; positive labeling for B72.3/TAG-72, monoclonal carcinoembryonic antigen, and mesothelin; and loss of DPC4 labeling can help distinguish metastatic pancreatic adenocarcinoma in the liver from bile duct adenomas or hamartomas. They also found that p63 and α-methylacyl coenzyme A racemase were specific for pancreatic adenocarcinoma, but the low sensitivity of these markers may limit their value in the clinical setting. Molecular analyses such as KRAS gene status may help in more difficult cases. For example, KRAS gene mutations are present in more than 90% of metastatic cancers but occur in less than 7% of benign bile duct proliferations.34 Perhaps more problematic is the distinction between a metastatic pancreatic cancer and an intrahepatic cholangiocarcinoma. Intrahepatic cholangiocarcinoma is most commonly seen as a well- to moderately differentiated adenocarcinoma forming small tubular glands in association with a prominent desmoplastic response, an appearance not unlike pancreatic cancer (Figure 4).35 However, Argani et al36 found that intrahepatic cholangiocarcinomas uncommonly show p53 and DPC4 loss by immunolabeling compared with distal common bile duct and pancreatic cancers, providing an avenue to aid correct diagnosis of these lesions.

Figure 3.

Pathology of bile duct adenomas and bile duct hamartomas. A, In this example of a bile duct adenoma, a haphazard arrangement of small tubules is seen that is admixed with fibrous stroma (hematoxylin-eosin, original magnification ×200). B, At higher power, the bland appearance of the epithelium and lack of mitotic figures is evident (hematoxylin-eosin, original magnification ×400). C, Bile duct hamartoma (von Meyenburg complex) showing dilated bile ducts containing refractile green-brown bile. Note the prominence of dense fibrotic stroma relative to the bile duct adenoma shown in A (hematoxylin-eosin, original magnification ×40). D, At higher power, the dilated bile ducts are lined by low cuboidal to flat epithelium with small, regular nuclei. The arrowhead indicates bile within a cystic space (hematoxylin-eosin, original magnification ×400)

Figure 3.

Pathology of bile duct adenomas and bile duct hamartomas. A, In this example of a bile duct adenoma, a haphazard arrangement of small tubules is seen that is admixed with fibrous stroma (hematoxylin-eosin, original magnification ×200). B, At higher power, the bland appearance of the epithelium and lack of mitotic figures is evident (hematoxylin-eosin, original magnification ×400). C, Bile duct hamartoma (von Meyenburg complex) showing dilated bile ducts containing refractile green-brown bile. Note the prominence of dense fibrotic stroma relative to the bile duct adenoma shown in A (hematoxylin-eosin, original magnification ×40). D, At higher power, the dilated bile ducts are lined by low cuboidal to flat epithelium with small, regular nuclei. The arrowhead indicates bile within a cystic space (hematoxylin-eosin, original magnification ×400)

Figure 4.

Intrahepatic cholangiocarcinoma. This moderately to poorly differentiated carcinoma is characterized by poorly formed glands, single infiltrating cells, and marked nuclear pleomorphism. In the absence of radiographic studies, clinical history, or immunolabeling, the histologic distinction from a metastatic pancreatic cancer would be exceedingly difficult (hematoxylin-eosin, original magnification ×400)

Figure 4.

Intrahepatic cholangiocarcinoma. This moderately to poorly differentiated carcinoma is characterized by poorly formed glands, single infiltrating cells, and marked nuclear pleomorphism. In the absence of radiographic studies, clinical history, or immunolabeling, the histologic distinction from a metastatic pancreatic cancer would be exceedingly difficult (hematoxylin-eosin, original magnification ×400)

Metastatic pancreatic adenocarcinoma to the lung can be difficult to distinguish from primary lung cancer, a situation compounded by the fact that both tumor types have a known association with smoking.37,38 Karanjawala et al39 reported that metastatic pancreatic adenocarcinomas in the lung commonly grow along the alveoli (lepidic growth) and display a mucinous-type epithelium, thus mimicking primary mucinous bronchioloalveolar carcinoma (Figure 5). Of interest, in 20% of cases no evidence of a desmoplastic response was found even though prominent desmoplasia was a feature of the primary carcinoma and/or metastatic deposits in other organ sites. Negative immunohistochemical labeling for thyroid transcription factor 1 in association with loss of DPC4 labeling was helpful to distinguish primary lung cancer from metastatic pancreatic adenocarcinoma to the lung. However, loss of DPC4 was the only reliable marker because mucinous bronchioloalveolar carcinomas were frequently negative for thyroid transcription factor 1.39 

Figure 5.

Histologic features of metastatic pancreatic carcinoma to the lung. A, The appearance and lepidic growth pattern of this metastatic pancreatic cancer can be histologically indistinguishable from a primary mucinous bronchioloalveolar carcinoma of the lung (hematoxylin-eosin, original magnification ×200). Compare the histologic features of this lung metastasis with the primary pancreatic cancer in the same patient shown in B (hematoxylin-eosin, original magnification ×200). C, The pancreatic origin of the lung metastasis shown in A was confirmed by loss of DPC4 immunolabeling (original magnification ×200). D, A similar pattern (DPC4 loss) is seen in the corresponding primary carcinoma (original magnification ×200)

Figure 5.

Histologic features of metastatic pancreatic carcinoma to the lung. A, The appearance and lepidic growth pattern of this metastatic pancreatic cancer can be histologically indistinguishable from a primary mucinous bronchioloalveolar carcinoma of the lung (hematoxylin-eosin, original magnification ×200). Compare the histologic features of this lung metastasis with the primary pancreatic cancer in the same patient shown in B (hematoxylin-eosin, original magnification ×200). C, The pancreatic origin of the lung metastasis shown in A was confirmed by loss of DPC4 immunolabeling (original magnification ×200). D, A similar pattern (DPC4 loss) is seen in the corresponding primary carcinoma (original magnification ×200)

The distinction of metastatic pancreatic carcinomas in the ovary from primary ovarian mucinous tumors (atypical proliferative/borderline and carcinoma) can also be difficult because of similarities in morphology.40 Pancreatic carcinomas are notorious for simulating primary ovarian mucinous carcinomas by displaying a multicystic appearance with relatively bland mucinous epithelium simulating a mucinous cystadenoma or an atypical proliferative (borderline) mucinous tumor41 (Figure 6). Seidman et al41 proposed the simple rule that classifies all bilateral mucinous carcinomas as metastasis, unilateral mucinous carcinoma less than 10 cm as metastatic, and unilateral mucinous carcinomas more than 10 cm as primary. However, metastatic pancreatic carcinomas are an exception in that they have a tendency for large growth and cystic change. Thus, features that suggest metastatic pancreatic cancer specifically rather than an ovarian primary include bilaterality, surface desmoplastic implants, size more than 10 cm, and extraovarian spread. Similar to other target organ sites, routine immunolabeling can also aid in diagnosis. Ji et al42 reported that loss of DPC4 labeling is useful when used in an immunolabeling panel for distinguishing metastatic pancreatic carcinomas in the ovary from both primary ovarian mucinous tumors and metastatic mucinous carcinomas derived from other sites. More recently, Cao et al43 found that an immunohistochemical panel including DPC4, mesothelin, fascin, and prostate stem cell antigen is useful for evaluating difficult mucinous tumors in the ovary when the differential diagnosis includes metastatic pancreatic adenocarcinoma.

Figure 6.

Histologic features of metastatic pancreatic carcinoma to the ovary. A, The multicystic appearance of this metastatic pancreatic cancer coupled with relatively bland mucinous glands simulates a primary ovarian mucinous tumor (hematoxylin-eosin, original magnification ×200). B, Immunolabeling for DPC4 supports the pancreatic origin of this metastatic neoplasm (original magnification ×200)

Figure 6.

Histologic features of metastatic pancreatic carcinoma to the ovary. A, The multicystic appearance of this metastatic pancreatic cancer coupled with relatively bland mucinous glands simulates a primary ovarian mucinous tumor (hematoxylin-eosin, original magnification ×200). B, Immunolabeling for DPC4 supports the pancreatic origin of this metastatic neoplasm (original magnification ×200)

GENETICS

The last decade has seen a dramatic increase in our understanding of the molecular biology of pancreatic cancer.5 Genome-based studies of pancreatic cancer have identified high rates of allelic loss of chromosomes 9p, 18q, 17p, 1p, and 6q in a series of primary pancreatic cancer xenografts.44,45 The targets of these losses include the tumor suppressor genes p16/CDKN2A on chromosome 9p,46 ,DPC4 (SMAD4, MADH4) on chromosome 18q,47 ,TP53 and MKK4 on chromosome 17p,48,49 and BRCA2 on chromosome 13q.50 The identification of noninvasive precursor lesions of invasive pancreatic ductal adenocarcinoma, pancreatic intraepithelial neoplasia, represents a significant advance in our understanding of the earliest forms of this disease.51 The genetic alterations identified in primary pancreatic cancer and its precursors are now thought to follow a genetic progression model associated with the accumulation of multiple genetic alterations, including telomere shortening52; activating point mutations in the KRAS2 gene; and inactivation of the p16/CDKN2A, TP53, SMAD4/DPC4, and BRCA2 tumor suppressor genes51 (Figure 7). Additional genes targeted in pancreatic cancer have been identified using high-resolution global analysis technologies.53–57 For example, novel candidate oncogenes include SMURF1 on 7q21, FGFR1 on 8p12, BIRC2 and BIRC3 on 11q22, and PAK4 on 19q13, whereas novel candidate tumor suppressor genes include TUSC3 on 8p22 and FEZ1 on 8p23.54–58 

Figure 7.

Genetic progression model of pancreatic carcinogenesis. The progression from histologically normal epithelium to low-grade pancreatic intraepithelial neoplasia (PanIN) (PanIN-1 and PanIN-2), to high-grade PanIN-3, to invasive carcinoma (left to right) is associated with the accumulation of specific genetic alterations. On the basis of their temporal appearance in this progression model, the molecular abnormalities can be classified as early (KRAS2 mutation, telomere shortening), intermediate (p16/CDKN2A loss), or late (mutations of DPC4/SMAD4, TP53, BRCA2). However, those changes that specifically promote progression to metastatic disease remain unknown

Figure 7.

Genetic progression model of pancreatic carcinogenesis. The progression from histologically normal epithelium to low-grade pancreatic intraepithelial neoplasia (PanIN) (PanIN-1 and PanIN-2), to high-grade PanIN-3, to invasive carcinoma (left to right) is associated with the accumulation of specific genetic alterations. On the basis of their temporal appearance in this progression model, the molecular abnormalities can be classified as early (KRAS2 mutation, telomere shortening), intermediate (p16/CDKN2A loss), or late (mutations of DPC4/SMAD4, TP53, BRCA2). However, those changes that specifically promote progression to metastatic disease remain unknown

Most major molecular studies of pancreatic cancer have only used tissues collected from resection specimens8,51,59–61; thus, the molecular features of advanced stage disease are relatively unknown. Nonetheless, in those studies that have focused on advanced stage pancreatic cancer, differences have been found indicating there is additional information to be gleaned.

KRAS2

Activation of the KRAS2 oncogene follows point mutation of codon 12 and, less often, of codons 13 and 61. These mutations maintain the protein in its active form independent of external signals resulting in activation of downstream effector pathways such as Raf-1, Rac, Rho, or phospho-inositol-3-kinase.62 ,KRAS2 gene mutations are among the most common mutations in pancreatic cancer and can be found in up to 95% of pancreatic carcinoma tissues.63,64 ,KRAS2 mutations can be found in early pancreatic intraepithelial neoplasia (pancreatic intraepithelial neoplasia 1), and the rate increases with advancing cellular atypia in precursor lesions.65,66 ,KRAS2 mutations seem to be prerequisite for pancreatic carcinogenesis but not metastasis, as these mutations are present universally in both primary and metastatic cancers in the same patients.11 

TP53

TP53 is the most commonly mutated tumor suppressor gene in all human cancers.67 It is a nuclear phosphoprotein with the ability to bind to specific DNA elements and to activate gene transcription. p53 has a central position in cell cycle regulation through its role in inactivating a variety of genes and interrupting cell proliferation at the G1-S checkpoint. Inactivating mutations in TP53 are found in up to 70% of pancreatic cancers, most often due to loss of heterozygosity coupled with a mutation in the remaining allele.48,68,69 In one of the few studies to assess TP53 status in metastatic pancreatic cancer, Ruggeri et al68 immunohistochemically analyzed p53 expression in primary and metastatic pancreatic cancer and found no difference in the rate of abnormal p53 labeling between each group. Their study also included a comparison of p53 labeling in 17 matched pairs of primary cancer and metastasis that indicated a strong correlation of abnormal labeling between the primary and metastatic sites. By contrast, no correlation was found with tumor grade, stage, or metastatic disease. More recently, Embuscado et al evaluated TP53 in a series of metastatic pancreatic cancer tissues obtained at autopsy and also found that TP53 showed a similar rate of inactivation to that reported in early stage pancreatic cancers.11,48 Overall, these 2 studies lend support to the hypothesis that genetic alteration of TP53 occurs relatively early in pancreatic carcinogenesis prior to the development of metastatic spread.

DPC4

DPC4 (SMAD4, MADH4) is a tumor suppressor gene that encodes for a critical transcription factor involved in the TGF-β signal pathway. Genetic inactivation of the DPC4 gene occurs in 55% of pancreatic adenocarcinomas, most commonly by homozygous deletions or intragenic mutations combined with loss of heterozygosity.47 The TGF-β signaling pathway is known to induce epithelial-mesenchymal transition,25–27 an event that occurs normally during embryonic development and is thought to be a critical event contributing to tumor invasiveness and metastasis.70 Consistent with this notion, there is a significantly greater frequency of DPC4 loss in metastatic pancreatic cancers compared with early stage cancers or pancreatic intraepithelial neoplasias, and loss of DPC4 protein expression has been shown to correlate with a shortened survival in patients with resectable pancreatic cancers.11,71–73 The mechanistic relationship of DPC4 loss and/or TGF-β signaling dysregulation to metastatic propensity of pancreatic cancer is uncertain but may relate to TGF-β–mediated signaling through the PI3K/PTEN pathway.25–27 

MKK4

MKK4 (MP2K4/SEK1) is a member of the mitogen-activated protein kinase family, originally identified as a kinase involved in the stress-activated protein kinase pathway by directly phosphorylating c-Jun NH2-terminal kinase.74 ,MKK4 genetic inactivation by homozygous deletion or mutation has been observed in a minority of pancreatic carcinomas, implicating deregulation of the stress-activated protein kinase pathway in pancreatic carcinogenesis.49 Xin et al75 evaluated MKK4 protein expression patterns by immunohistochemical labeling in a large series of resected, advanced stage, and metastatic pancreatic cancers and noted that loss of MKK4 immunolabeling was significantly more common in distant metastases than in the primary carcinomas in these same patients. Moreover, among matched primary and metastatic cancer tissues, loss of MKK4 immunolabeling was seen in 11% of primary cancers but in 37% of distant metastases indicating that MKK4 loss correlates with metastasis formation. The MKK4 gene has been reported to function as a metastasis-suppressor gene in ovarian and prostatic carcinomas, in which the loss of MKK4 gene expression facilitates metastasis formation without affecting primary tumor growth.76 However, using a somatic gene knockout of MKK4 in human cancer cells, Cunningham et al77,78 found that the MKK4 null phenotype was markedly deficient in producing experimental metastases compared with wild-type pancreatic cancer cells and rationalized that a gene dosage-dependent phenotype of MKK4 was needed for optimal metastatic spread of pancreatic cancer.

EphA2

EphA2 is a transmembrane receptor tyrosine kinase that functions in the regulation of cell growth, survival, angiogenesis, and migration.79 EphA2 is overexpressed in a variety of human cancers, and high levels of EphA2 are associated with aggressive disease and poor clinical outcomes.80 In pancreatic cancers, EphA2 protein is also relatively overexpressed in pancreatic adenocarcinoma cell lines with higher metastatic potential and suppression of EphA2 expression appears to attenuate the invasive phenotype of these same cell lines.81,82 

Mudali et al83 evaluated the immunohistochemical labeling patterns of EphA2 protein in a large series of matched primary and metastatic pancreatic cancers and also found frequent overexpression of EphA2 in pancreatic carcinomas compared with benign ducts or pancreatic intraepithelial neoplasia lesions. However, they also noted a significant correlation of EphA2 immunolabeling intensity and organ site metastasis, in that relatively low levels of EphA2 labeling were seen in liver metastases, whereas high levels were seen in association with lymph node metastases (Figure 8). The authors hypothesized that in addition to promoting metastasis, EphA2 levels may contribute to organ site–specific metastasis as well. In rare cases, overexpression of EphA2 was due to amplification at chromosome 1p36, providing a potentially genetic basis for enhanced metastatic ability. These findings are exciting as antibody-mediated targeting of EphA2 has been shown to cause EphA2 internalization, degradation, and loss of oncogenic signals, thus highlighting EphA2 as a therapeutic target for advanced stage pancreatic cancer.84 

Figure 8.

EphA2 labeling in pancreatic cancer and matched metastases to liver and lymph node. The primary carcinoma in A shows positive labeling for EphA2. By contrast, intense positive labeling is seen in the lymph node metastasis (B), whereas virtually negative labeling is seen for the liver metastasis from the same patient (C) (original magnifications ×400)

Figure 8.

EphA2 labeling in pancreatic cancer and matched metastases to liver and lymph node. The primary carcinoma in A shows positive labeling for EphA2. By contrast, intense positive labeling is seen in the lymph node metastasis (B), whereas virtually negative labeling is seen for the liver metastasis from the same patient (C) (original magnifications ×400)

Gene Expression Profiling

Global expression platforms, such as complementary DNA or oligonucleotide microarrays and serial analysis of gene expression, have helped elucidate large numbers of differentially expressed transcripts in pancreatic cancer. However, most studies have largely focused on the differential gene expression between normal pancreatic tissue and primary pancreatic adenocarcinoma and thus have not addressed pancreatic cancer progression and metastasis specifically.61,85–89 

Campagna et al90 used gene expression microarrays to evaluate human pancreatic cancer tissues for global changes in expression that accompany metastatic spread and found no reproducible differences between primary cancers and metastases, whether using matched or unmatched samples. However, a comparison of the expression profiles of early stage and late stage primary carcinomas specifically revealed significant differences in gene expression that were most prominent in gene classes that related to the MAPK and Wnt pathways, metabolism, immune regulation, and cell-cell and cell-matrix interactions. Of interest, one of the genes identified and validated was MAX interacting protein 1 (MXI1), a member of the max dimerization protein family of transcriptional repressors that negatively regulates c-myc.91 Amplification and overexpression of c-myc has been well described in pancreatic cancer where it acts as a central regulator of gene transcription and predisposes cells to apoptosis under nutrient, growth factor, or oxygen deprivation conditions.92,93 Thus, although c-myc amplification and/or overexpression may promote pancreatic carcinogenesis,94 upregulation of MXI1 may occur during disease progression to refine c-myc oncogenic signals and to protect cancer cells from c-myc–induced apoptosis.93 This expression profile associated with growth at the primary site did not overlap with that found when comparing normal pancreatic tissues to pancreatic cancers, leading Campagna et al90 to suggest that in addition to the robust changes in expression that accompany pancreatic carcinogenesis, specific and focused changes occur in association with growth at the primary site that may promote the outgrowth of metastatic clones.

Due to the lack of available high-quality metastatic cancer tissues for expression profiling, a variety of studies have addressed pancreatic cancer metastasis using in vivo models of spontaneous liver metastasis from orthotopic xenografts.95–98 Although all have demonstrated significant differences in gene expression among the primary and metastatic sites, there is little overlap among different gene sets to suggest a commonly dysregulated pathway in pancreatic cancer metastasis formation. For example, Niedergethmann et al98 performed transcriptional profiling of orthotopic xenografts and identified 25 statistically significant upregulated and 181 downregulated genes when comparing the liver metastases with the primary orthotopic tumor. Eight of these genes (PAI-1, BNIP3l, VEGF, NSE, RGS4, HSP27, GADD45A, PTPN14) were validated in a semiquantitative immunohistochemical analysis, and overexpression analyses revealed 66 significantly dysregulated pathways associated with cell proliferation, cell stress, cell communication, and metabolic and cytokine function. Suemizu et al96 also identified statistically significant differences in gene expression between orthotopic xenografts and their spontaneous metastases. However, in their study, only S100A4 expression correlated with the ability to form liver metastases, as evaluated in a quantitative model of metastasis in NOG (NOD/Shi-scid IL2r γ null) mice.

Nomura et al99 described a robust system in which they established HPC-1H5, a highly liver metastatic cell line, and HPC-1P5a, a highly peritoneal disseminating cell line, from the parental pancreatic cancer cell line HPC-1. Using expression profiling to interrogate the gene expression of each line, they found nonoverlapping sets of dysregulated genes in the hepatic and peritoneal disseminating cell lines. For example, several integrins such as α1 integrin (CD49A), α2 integrin (CD49B), α2b integrin (CD41), and β1 integrin (CD29) were upregulated in the liver metastasizing cell line, whereas angiopoetin 1 and MMP10 were upregulated genes of the peritoneal disseminating cell line.

Hedgehog Signaling Pathway

Hedgehog expression is normally restricted to embryogenesis and is instrumental in patterning tissues and structures of the early embryo, including the pancreas.100,101 After pancreatic development, Hedgehog signaling is restricted to the maintenance of stem cell pools.102,103 In normal Hedgehog pathway function, the transporter-like protein Patched acts catalytically to inhibit activation of the 7 transmembrane protein Smoothened. Patched activity is blocked by binding of Hedgehog ligand, thus liberating Smoothened for activation of transcriptional targets mediated by the Gli family of latent transcriptional factors (Figure 9).103 

Figure 9.

Simple representation of Hedgehog (Hh) signaling in normal cells.103 Activation of the Hh signaling pathway is initiated by binding of a Hh ligand to Patched (Ptch). This lifts suppression by Ptch of Smoothened (Smo), activating a cascade that leads to the nuclear translocation of Gli and the activation of target genes. A variety of Gli interacting proteins (indicated by orange) such as Rab23, FKBP8, and the intraflagellar transport proteins act downstream of Ptch and Smo to regulate Gli activity. Gli and Ptch are also target genes of Gli, thus perpetuating a feedback loop of Hh signaling. Hh-interacting protein (HIP) is a membrane-bound protein that antagonizes Hh pathway activity by binding to Hh ligands

Figure 9.

Simple representation of Hedgehog (Hh) signaling in normal cells.103 Activation of the Hh signaling pathway is initiated by binding of a Hh ligand to Patched (Ptch). This lifts suppression by Ptch of Smoothened (Smo), activating a cascade that leads to the nuclear translocation of Gli and the activation of target genes. A variety of Gli interacting proteins (indicated by orange) such as Rab23, FKBP8, and the intraflagellar transport proteins act downstream of Ptch and Smo to regulate Gli activity. Gli and Ptch are also target genes of Gli, thus perpetuating a feedback loop of Hh signaling. Hh-interacting protein (HIP) is a membrane-bound protein that antagonizes Hh pathway activity by binding to Hh ligands

Aberrant reactivation of the Hedgehog signaling pathways, and concomitant overexpression of its target genes, is a common feature of pancreatic cancer.104,105 Hedgehog signaling has also been implicated in pancreatic cancer metastasis. Feldmann et al106 reported that Hedgehog target gene GLI1 levels are upregulated in tissue samples of metastatic human pancreatic cancer samples compared with matched primary tumors. Moreover, inhibition of Hedgehog signaling with the small molecular cyclopamine caused a profound decrease in metastatic spread in this orthotopic xenograft model. A combination of gemcitabine and cyclopamine completely abrogated metastases in this same model while significantly reducing the size of the orthotopic tumors, suggesting a central role of the Hedgehog pathway in pancreatic cancer metastasis.

SUMMARY

Metastatic spread from pancreatic cancer remains a major hurdle in combating this disease. A better understanding of the clinical, genetic, and molecular features of metastatic pancreatic cancer will afford novel areas for therapeutic intervention.

References

References
Jemal
,
A.
,
R.
Siegel
, and
E.
Ward
.
et al
.
Cancer statistics, 2008.
CA Cancer J Clin
2008
.
58
:
71
96
.
Sohn
,
T. A.
,
C. J.
Yeo
, and
J. L.
Cameron
.
et al
.
Resected adenocarcinoma of the pancreas—616 patients: results, outcomes, and prognostic indicators.
J Gastrointest Surg
2000
.
4
:
567
579
.
Winter
,
J. M.
,
J. L.
Cameron
, and
K. A.
Campbell
.
et al
.
1423 pancreaticoduodenectomies for pancreatic cancer: a single-institution experience.
J Gastrointest Surg
2006
.
10
:
1199
1210
.
discussion 1210–1211
.
Moore
,
M. J.
,
D.
Goldstein
, and
J.
Hamm
.
et al
.
Erlotinib plus gemcitabine compared with gemcitabine alone in patients with advanced pancreatic cancer: a phase III trial of the National Cancer Institute of Canada clinical trials group.
J Clin Oncol
2007
.
25
:
1960
1966
.
Maitra
,
A.
and
R. H.
Hruban
.
Pancreatic cancer.
Annu Rev Pathol
2008
.
3
:
157
188
.
Vogelstein
,
B.
and
K. W.
Kinzler
.
Cancer genes and the pathways they control.
Nat Med
2004
.
10
:
789
799
.
Pawlik
,
T. M.
and
M. A.
Choti
.
Surgical therapy for colorectal metastases to the liver.
J Gastrointest Surg
2007
.
11
:
1057
1077
.
Verslype
,
C.
,
E.
Van Cutsem
, and
M.
Dicato
.
et al
.
The management of pancreatic cancer: current expert opinion and recommendations derived from the 8th World Congress on Gastrointestinal Cancer, Barcelona, 2006.
Ann Oncol
2007
.
18
:(
suppl 7
).
vii1
vii10
.
Kamisawa
,
T.
,
T.
Isawa
,
M.
Koike
,
K.
Tsuruta
, and
A.
Okamoto
.
Hematogenous metastases of pancreatic ductal carcinoma.
Pancreas
1995
.
11
:
345
349
.
Mao
,
C.
,
D. R.
Domenico
,
K.
Kim
,
D. J.
Hanson
, and
J. M.
Howard
.
Observations on the developmental patterns and the consequences of pancreatic exocrine adenocarcinoma: findings of 154 autopsies.
Arch Surg
1995
.
130
:
125
134
.
Embuscado
,
E. E.
,
D.
Laheru
, and
F.
Ricci
.
et al
.
Immortalizing the complexity of cancer metastasis: genetic features of lethal metastatic pancreatic cancer obtained from rapid autopsy.
Cancer Biol Ther
2005
.
4
:
548
554
.
Disibio
,
G.
and
S. W.
French
.
Metastatic patterns of cancers: results from a large autopsy study.
Arch Pathol Lab Med
2008
.
132
:
931
939
.
Amikura
,
K.
,
M.
Kobari
, and
S.
Matsuno
.
The time of occurrence of liver metastasis in carcinoma of the pancreas.
Int J Pancreatol
1995
.
17
:
139
146
.
Vogel
,
I.
,
U.
Kruger
, and
J.
Marxsen
.
et al
.
Disseminated tumor cells in pancreatic cancer patients detected by immunocytology: a new prognostic factor.
Clin Cancer Res
1999
.
5
:
593
599
.
Roder
,
J. D.
,
S.
Thorban
,
K.
Pantel
, and
J. R.
Siewert
.
Micrometastases in bone marrow: prognostic indicators for pancreatic cancer.
World J Surg
1999
.
23
:
888
891
.
Yachida
,
S.
,
N.
Fukushima
,
M.
Sakamoto
,
Y.
Matsuno
,
T.
Kosuge
, and
S.
Hirohashi
.
Implications of peritoneal washing cytology in patients with potentially resectable pancreatic cancer.
Br J Surg
2002
.
89
:
573
578
.
Nakahashi
,
C.
,
T.
Oda
, and
T.
Kinoshita
.
et al
.
The impact of liver metastasis on mortality in patients initially diagnosed with locally advanced or resectable pancreatic cancer.
Int J Gastrointest Cancer
2003
.
33
:
155
164
.
Hishinuma
,
S.
,
Y.
Ogata
,
J.
Matsui
, and
I.
Ozawa
.
Results of surgery and adjuvant radiotherapy for pancreatic cancer.
J Hepatobiliary Pancreat Surg
1998
.
5
:
143
150
.
Iacobuzio-Donahue
,
C. A.
,
B.
Fu
, and
S.
Yachida
.
et al
.
Dpc4 gene status of the primary carcinoma correlates with local versus metastatic failure in patients with pancreatic cancer.
J Clin Oncol. 2009. In press
.
Kayahara
,
M.
,
T.
Nagakawa
,
K.
Ueno
,
T.
Ohta
,
T.
Takeda
, and
I.
Miyazaki
.
An evaluation of radical resection for pancreatic cancer based on the mode of recurrence as determined by autopsy and diagnostic imaging.
Cancer
1993
.
72
:
2118
2123
.
Greer
,
S. E.
,
J. M.
Pipas
, and
J. E.
Sutton
.
et al
.
Effect of neoadjuvant therapy on local recurrence after resection of pancreatic adenocarcinoma.
J Am Coll Surg
2008
.
206
:
451
457
.
Farnell
,
M. B.
,
R. K.
Pearson
, and
M. G.
Sarr
.
et al
.
A prospective randomized trial comparing standard pancreatoduodenectomy with pancreatoduodenectomy with extended lymphadenectomy in resectable pancreatic head adenocarcinoma.
Surgery
2005
.
138
:
618
628
.
discussion 628–630
.
Yeo
,
C. J.
,
J. L.
Cameron
, and
K. D.
Lillemoe
.
et al
.
Pancreaticoduodenectomy with or without distal gastrectomy and extended retroperitoneal lymphadenectomy for periampullary adenocarcinoma, part 2: randomized controlled trial evaluating survival, morbidity, and mortality.
Ann Surg
2002
.
236
:
355
366
.
discussion 366– 358
.
Winter
,
J. M.
,
A. H.
Ting
, and
F.
Vilardell
.
et al
.
Absence of e-cadherin expression distinguishes noncohesive from cohesive pancreatic cancer.
Clin Cancer Res
2008
.
14
:
412
418
.
Derynck
,
R.
and
R. J.
Akhurst
.
Differentiation plasticity regulated by TGF-beta family proteins in development and disease.
Nat Cell Biol
2007
.
9
:
1000
1004
.
Vogelmann
,
R.
,
M. D.
Nguyen-Tat
,
K.
Giehl
,
G.
Adler
,
D.
Wedlich
, and
A.
Menke
.
TGFbeta-induced downregulation of E-cadherin-based cell-cell adhesion depends on PI3-kinase and PTEN.
J Cell Sci
2005
.
118
:
4901
4912
.
Deckers
,
M.
,
M.
van Dinther
, and
J.
Buijs
.
et al
.
The tumor suppressor smad4 is required for transforming growth factor beta-induced epithelial to mesenchymal transition and bone metastasis of breast cancer cells.
Cancer Res
2006
.
66
:
2202
2209
.
Terayama
,
N.
,
T.
Terada
, and
Y.
Nakanuma
.
Histologic growth patterns of metastatic carcinomas of the liver.
Jpn J Clin Oncol
1996
.
26
:
24
29
.
Yamamoto
,
J.
,
A.
Saiura
, and
R.
Koga
.
et al
.
Surgical treatment for metastatic malignancies: nonanatomical resection of liver metastasis: indications and outcomes.
Int J Clin Oncol
2005
.
10
:
97
102
.
Allaire
,
G. S.
,
L.
Rabin
,
K. G.
Ishak
, and
I. A.
Sesterhenn
.
Bile duct adenoma: a study of 152 cases.
Am J Surg Pathol
1988
.
12
:
708
715
.
Govindarajan
,
S.
and
R. L.
Peters
.
The bile duct adenoma: a lesion distinct from Meyenburg complex.
Arch Pathol Lab Med
1984
.
108
:
922
924
.
Shah
,
J. P.
,
H. S.
Goldsmith
, and
A. G.
Huvos
.
Hamartomas of the liver.
Surgery
1970
.
68
:
778
782
.
Hornick
,
J. L.
,
G. Y.
Lauwers
, and
R. D.
Odze
.
Immunohistochemistry can help distinguish metastatic pancreatic adenocarcinomas from bile duct adenomas and hamartomas of the liver.
Am J Surg Pathol
2005
.
29
:
381
389
.
Hruban
,
R. H.
,
P. D.
Sturm
, and
R. J.
Slebos
.
et al
.
Can k-ras codon 12 mutations be used to distinguish benign bile duct proliferations from metastases in the liver?: a molecular analysis of 101 liver lesions from 93 patients.
Am J Pathol
1997
.
151
:
943
949
.
Blechacz
,
B.
and
G. J.
Gores
.
Cholangiocarcinoma: advances in pathogenesis, diagnosis, and treatment.
Hepatology
2008
.
48
:
308
321
.
Argani
,
P.
,
A.
Shaukat
, and
M.
Kaushal
.
et al
.
Differing rates of loss of dpc4 expression and of p53 overexpression among carcinomas of the proximal and distal bile ducts.
Cancer
2001
.
91
:
1332
1341
.
Iodice
,
S.
,
S.
Gandini
,
P.
Maisonneuve
, and
A. B.
Lowenfels
.
Tobacco and the risk of pancreatic cancer: a review and meta-analysis.
Langenbecks Arch Surg
2008
.
393
:
535
545
.
Freedman
,
N. D.
,
M. F.
Leitzmann
,
A. R.
Hollenbeck
,
A.
Schatzkin
, and
C. C.
Abnet
.
Cigarette smoking and subsequent risk of lung cancer in men and women: analysis of a prospective cohort study.
Lancet Oncol
2008
.
9
:
649
656
.
Karanjawala
,
Z. E.
,
R. H.
Hruban
, and
C. A.
Iacobuzio-Donahue
.
Metastatic pancreatic cancer to the lung: a potential mimicker of primary bronchioalveolar carcinoma of the lung.
Mod Pathol
2008
.
21
:
307A
.
Young
,
R. H.
From Krukenberg to today: the ever present problems posed by metastatic tumors in the ovary: part II.
Adv Anat Pathol
2007
.
14
:
149
177
.
Seidman
,
J. D.
,
R. J.
Kurman
, and
B. M.
Ronnett
.
Primary and metastatic mucinous adenocarcinomas in the ovaries: incidence in routine practice with a new approach to improve intraoperative diagnosis.
Am J Surg Pathol
2003
.
27
:
985
993
.
Ji
,
H.
,
C.
Isacson
,
J. D.
Seidman
,
R. J.
Kurman
, and
B. M.
Ronnett
.
Cytokeratins 7 and 20, dpc4, and muc5ac in the distinction of metastatic mucinous carcinomas in the ovary from primary ovarian mucinous tumors: dpc4 assists in identifying metastatic pancreatic carcinomas.
Int J Gynecol Pathol
2002
.
21
:
391
400
.
Cao
,
D.
,
H.
Ji
, and
B. M.
Ronnett
.
Expression of mesothelin, fascin, and prostate stem cell antigen in primary ovarian mucinous tumors and their utility in differentiating primary ovarian mucinous tumors from metastatic pancreatic mucinous carcinomas in the ovary.
Int J Gynecol Pathol
2005
.
24
:
67
72
.
Iacobuzio-Donahue
,
C. A.
,
M. S.
van der Heijden
, and
M. R.
Baumgartner
.
et al
.
Large-scale allelotype of pancreaticobiliary carcinoma provides quantitative estimates of genome-wide allelic loss.
Cancer Res
2004
.
64
:
871
875
.
Calhoun
,
E. S.
,
T.
Hucl
, and
E.
Gallmeier
.
et al
.
Identifying allelic loss and homozygous deletions in pancreatic cancer without matched normals using high-density single-nucleotide polymorphism arrays.
Cancer Res
2006
.
66
:
7920
7928
.
Caldas
,
C.
,
S. A.
Hahn
, and
L. T.
da Costa
.
et al
.
Frequent somatic mutations and homozygous deletions of the p16 (mts1) gene in pancreatic adenocarcinoma.
Nat Genet
1994
.
8
:
27
32
.
Hahn
,
S. A.
,
M.
Schutte
, and
A. T.
Hoque
.
et al
.
Dpc4, a candidate tumor suppressor gene at human chromosome 18q21.1.
Science
1996
.
271
:
350
353
.
Redston
,
M. S.
,
C.
Caldas
, and
A. B.
Seymour
.
et al
.
P53 mutations in pancreatic carcinoma and evidence of common involvement of homocopolymer tracts in DNA microdeletions.
Cancer Res
1994
.
54
:
3025
3033
.
Su
,
G. H.
,
W.
Hilgers
, and
M. C.
Shekher
.
et al
.
Alterations in pancreatic, biliary, and breast carcinomas support mkk4 as a genetically targeted tumor suppressor gene.
Cancer Res
1998
.
58
:
2339
2342
.
Goggins
,
M.
,
M.
Schutte
, and
J.
Lu
.
et al
.
Germline brca2 gene mutations in patients with apparently sporadic pancreatic carcinomas.
Cancer Res
1996
.
56
:
5360
5364
.
Hruban
,
R. H.
,
M.
Goggins
,
J.
Parsons
, and
S. E.
Kern
.
Progression model for pancreatic cancer.
Clin Cancer Res
2000
.
6
:
2969
2972
.
van Heek
,
N. T.
,
A. K.
Meeker
, and
S. E.
Kern
.
et al
.
Telomere shortening is nearly universal in pancreatic intraepithelial neoplasia.
Am J Pathol
2002
.
161
:
1541
1547
.
Aguirre
,
A. J.
,
C.
Brennan
, and
G.
Bailey
.
et al
.
High-resolution characterization of the pancreatic adenocarcinoma genome.
Proc Natl Acad Sci U S A
2004
.
101
:
9067
9072
.
Nowak
,
N. J.
,
D.
Gaile
, and
J. M.
Conroy
.
et al
.
Genome-wide aberrations in pancreatic adenocarcinoma.
Cancer Genet Cytogenet
2005
.
161
:
36
50
.
Bashyam
,
M. D.
,
R.
Bair
, and
Y. H.
Kim
.
et al
.
Array-based comparative genomic hybridization identifies localized DNA amplifications and homozygous deletions in pancreatic cancer.
Neoplasia
2005
.
7
:
556
562
.
Mahlamaki
,
E. H.
,
P.
Kauraniemi
,
O.
Monni
,
M.
Wolf
,
S.
Hautaniemi
, and
A.
Kallioniemi
.
High-resolution genomic and expression profiling reveals 105 putative amplification target genes in pancreatic cancer.
Neoplasia
2004
.
6
:
432
439
.
Heidenblad
,
M.
,
E. F.
Schoenmakers
, and
T.
Jonson
.
et al
.
Genome-wide array-based comparative genomic hybridization reveals multiple amplification targets and novel homozygous deletions in pancreatic carcinoma cell lines.
Cancer Res
2004
.
64
:
3052
3059
.
Heidenblad
,
M.
,
D.
Lindgren
, and
J. A.
Veltman
.
et al
.
Microarray analyses reveal strong influence of DNA copy number alterations on the transcriptional patterns in pancreatic cancer: implications for the interpretation of genomic amplifications.
Oncogene
2005
.
24
:
1794
1801
.
Hahn
,
S. A.
,
A. B.
Seymour
, and
A. T.
Hoque
.
et al
.
Allelotype of pancreatic adenocarcinoma using xenograft enrichment.
Cancer Res
1995
.
55
:
4670
4675
.
Li
,
C.
,
D. G.
Heidt
, and
P.
Dalerba
.
et al
.
Identification of pancreatic cancer stem cells.
Cancer Res
2007
.
67
:
1030
1037
.
Iacobuzio-Donahue
,
C. A.
,
R.
Ashfaq
, and
A.
Maitra
.
et al
.
Highly expressed genes in pancreatic ductal adenocarcinomas: a comprehensive characterization and comparison of the transcription profiles obtained from three major technologies.
Cancer Res
2003
.
63
:
8614
8622
.
Kranenburg
,
O.
The kras oncogene: past, present, and future.
Biochim Biophys Acta
2005
.
1756
:
81
82
.
Smit
,
V. T.
,
A. J.
Boot
,
A. M.
Smits
,
G. J.
Fleuren
,
C. J.
Cornelisse
, and
J. L.
Bos
.
Kras codon 12 mutations occur very frequently in pancreatic adenocarcinomas.
Nucleic Acids Res
1988
.
16
:
7773
7782
.
Hruban
,
R. H.
,
A. D.
van Mansfeld
, and
G. J.
Offerhaus
.
et al
.
K-ras oncogene activation in adenocarcinoma of the human pancreas: a study of 82 carcinomas using a combination of mutant-enriched polymerase chain reaction analysis and allele-specific oligonucleotide hybridization.
Am J Pathol
1993
.
143
:
545
554
.
Moskaluk
,
C. A.
,
R. H.
Hruban
, and
S. E.
Kern
.
P16 and k-ras gene mutations in the intraductal precursors of human pancreatic adenocarcinoma.
Cancer Res
1997
.
57
:
2140
2143
.
Luttges
,
J.
,
B.
Schlehe
,
M. A.
Menke
,
I.
Vogel
,
D.
Henne-Bruns
, and
G.
Kloppel
.
The k-ras mutation pattern in pancreatic ductal adenocarcinoma usually is identical to that in associated normal, hyperplastic, and metaplastic ductal epithelium.
Cancer
1999
.
85
:
1703
1710
.
Vogelstein
,
B.
,
D.
Lane
, and
A. J.
Levine
.
Surfing the p53 network.
Nature
2000
.
408
:
307
310
.
Ruggeri
,
B. A.
,
L.
Huang
, and
D.
Berger
.
et al
.
Molecular pathology of primary and metastatic ductal pancreatic lesions: analyses of mutations and expression of the p53, mdm-2, and p21/waf-1 genes in sporadic and familial lesions.
Cancer
1997
.
79
:
700
716
.
Weyrer
,
K.
,
H.
Feichtinger
, and
M.
Haun
.
et al
.
P53, ki-ras, and DNA ploidy in human pancreatic ductal adenocarcinomas.
Lab Invest
1996
.
74
:
279
289
.
Zavadil
,
J.
and
E. P.
Bottinger
.
TGF-beta and epithelial-to-mesenchymal transitions.
Oncogene
2005
.
24
:
5764
5774
.
Wilentz
,
R. E.
,
C. A.
Iacobuzio-Donahue
, and
P.
Argani
.
et al
.
Loss of expression of dpc4 in pancreatic intraepithelial neoplasia: evidence that dpc4 inactivation occurs late in neoplastic progression.
Cancer Res
2000
.
60
:
2002
2006
.
Tascilar
,
M.
,
H. G.
Skinner
, and
C.
Rosty
.
et al
.
The smad4 protein and prognosis of pancreatic ductal adenocarcinoma.
Clin Cancer Res
2001
.
7
:
4115
4121
.
Biankin
,
A. V.
,
A. L.
Morey
, and
C. S.
Lee
.
et al
.
Dpc4/smad4 expression and outcome in pancreatic ductal adenocarcinoma.
J Clin Oncol
2002
.
20
:
4531
4542
.
Sanchez
,
I.
,
R. T.
Hughes
, and
B. J.
Mayer
.
et al
.
Role of sapk/erk kinase-1 in the stress-activated pathway regulating transcription factor c-jun.
Nature
1994
.
372
:
794
798
.
Xin
,
W.
,
K. J.
Yun
, and
F.
Ricci
.
et al
.
Map2k4/mkk4 expression in pancreatic cancer: genetic validation of immunohistochemistry and relationship to disease course.
Clin Cancer Res
2004
.
10
:
8516
8520
.
Yamada
,
S. D.
,
J. A.
Hickson
, and
Y.
Hrobowski
.
et al
.
Mitogen-activated protein kinase kinase 4 (mkk4) acts as a metastasis suppressor gene in human ovarian carcinoma.
Cancer Res
2002
.
62
:
6717
6723
.
Cunningham
,
S. C.
,
E.
Gallmeier
, and
T.
Hucl
.
et al
.
Targeted deletion of mkk4 in cancer cells: a detrimental phenotype manifests as decreased experimental metastasis and suggests a counterweight to the evolution of tumor-suppressor loss.
Cancer Res
2006
.
66
:
5560
5564
.
Cunningham
,
S. C.
,
E.
Gallmeier
, and
T.
Hucl
.
et al
.
Theoretical proposal: allele dosage of map2k4/mkk4 could rationalize frequent 17p loss in diverse human cancers.
Cell Cycle
2006
.
5
:
1090
1093
.
Pasquale
,
E. B.
Eph receptor signalling casts a wide net on cell behaviour.
Nat Rev Mol Cell Biol
2005
.
6
:
462
475
.
Walker-Daniels
,
J.
,
A. R.
Hess
,
M. J.
Hendrix
, and
M. S.
Kinch
.
Differential regulation of epha2 in normal and malignant cells.
Am J Pathol
2003
.
162
:
1037
1042
.
Duxbury
,
M. S.
,
H.
Ito
,
M. J.
Zinner
,
S. W.
Ashley
, and
E. E.
Whang
.
Epha2: a determinant of malignant cellular behavior and a potential therapeutic target in pancreatic adenocarcinoma.
Oncogene
2004
.
23
:
1448
1456
.
Duxbury
,
M. S.
,
H.
Ito
,
M. J.
Zinner
,
S. W.
Ashley
, and
E. E.
Whang
.
Ligation of epha2 by ephrin a1-fc inhibits pancreatic adenocarcinoma cellular invasiveness.
Biochem Biophys Res Commun
2004
.
320
:
1096
1102
.
Mudali
,
S. V.
,
B.
Fu
,
S. S.
Lakkur
,
M.
Luo
,
E. E.
Embuscado
, and
C. A.
Iacobuzio-Donahue
.
Patterns of epha2 protein expression in primary and metastatic pancreatic carcinoma and correlation with genetic status.
Clin Exp Metastasis
2006
.
23
:
357
365
.
Carles-Kinch
,
K.
,
K. E.
Kilpatrick
,
J. C.
Stewart
, and
M. S.
Kinch
.
Antibody targeting of the epha2 tyrosine kinase inhibits malignant cell behavior.
Cancer Res
2002
.
62
:
2840
2847
.
Missiaglia
,
E.
,
E.
Blaveri
, and
B.
Terris
.
et al
.
Analysis of gene expression in cancer cell lines identifies candidate markers for pancreatic tumorigenesis and metastasis.
Int J Cancer
2004
.
112
:
100
112
.
Ryu
,
B.
,
J.
Jones
, and
N. J.
Blades
.
et al
.
Relationships and differentially expressed genes among pancreatic cancers examined by large-scale serial analysis of gene expression.
Cancer Res
2002
.
62
:
819
826
.
Grutzmann
,
R.
,
C.
Pilarsky
, and
O.
Ammerpohl
.
et al
.
Gene expression profiling of microdissected pancreatic ductal carcinomas using high-density DNA microarrays.
Neoplasia
2004
.
6
:
611
622
.
Crnogorac-Jurcevic
,
T.
,
E.
Efthimiou
, and
T.
Nielsen
.
et al
.
Expression profiling of microdissected pancreatic adenocarcinomas.
Oncogene
2002
.
21
:
4587
4594
.
Logsdon
,
C. D.
,
D. M.
Simeone
, and
C.
Binkley
.
et al
.
Molecular profiling of pancreatic adenocarcinoma and chronic pancreatitis identifies multiple genes differentially regulated in pancreatic cancer.
Cancer Res
2003
.
63
:
2649
2657
.
Campagna
,
D.
,
L.
Cope
,
S. S.
Lakkur
,
C. M.
Henderson
,
D.
Laheru
, and
C. A.
Iacobuzio-Donahue
.
Gene expression profiles associated with advanced pancreatic cancer.
Int J Clin Exp Pathol
2008
.
1
:
32
43
.
Hurlin
,
P. J.
and
J.
Huang
.
The max-interacting transcription factor network.
Semin Cancer Biol
2006
.
16
:
265
274
.
Dang
,
C. V.
,
K. A.
O'Donnell
,
K. I.
Zeller
,
T.
Nguyen
,
R. C.
Osthus
, and
F.
Li
.
The c-myc target gene network.
Semin Cancer Biol
2006
.
16
:
253
264
.
Corn
,
P. G.
,
M. S.
Ricci
, and
K. A.
Scata
.
et al
.
Mxi1 is induced by hypoxia in a hif-1-dependent manner and protects cells from c-myc-induced apoptosis.
Cancer Biol Ther
2005
.
4
:
1285
1294
.
Schleger
,
C.
,
C.
Verbeke
,
R.
Hildenbrand
,
H.
Zentgraf
, and
U.
Bleyl
.
C-myc activation in primary and metastatic ductal adenocarcinoma of the pancreas: incidence, mechanisms, and clinical significance.
Mod Pathol
2002
.
15
:
462
469
.
Loukopoulos
,
P.
,
K.
Kanetaka
,
M.
Takamura
,
T.
Shibata
,
M.
Sakamoto
, and
S.
Hirohashi
.
Orthotopic transplantation models of pancreatic adenocarcinoma derived from cell lines and primary tumors and displaying varying metastatic activity.
Pancreas
2004
.
29
:
193
203
.
Suemizu
,
H.
,
M.
Monnai
,
Y.
Ohnishi
,
M.
Ito
,
N.
Tamaoki
, and
M.
Nakamura
.
Identification of a key molecular regulator of liver metastasis in human pancreatic carcinoma using a novel quantitative model of metastasis in nod/scid/gammacnull (nog) mice.
Int J Oncol
2007
.
31
:
741
751
.
Tarbe
,
N.
,
V.
Evtimova
,
H.
Burtscher
,
M.
Jarsch
,
F.
Alves
, and
U. H.
Weidle
.
Transcriptional profiling of cell lines derived from an orthotopic pancreatic tumor model reveals metastasis-associated genes.
Anticancer Res
2001
.
21
:
3221
3228
.
Niedergethmann
,
M.
,
F.
Alves
, and
J. K.
Neff
.
et al
.
Gene expression profiling of liver metastases and tumour invasion in pancreatic cancer using an orthotopic scid mouse model.
Br J Cancer
2007
.
97
:
1432
1440
.
Nomura
,
H.
,
H.
Nishimori
, and
T.
Yasoshima
.
et al
.
A new liver metastatic and peritoneal dissemination model established from the same human pancreatic cancer cell line: analysis using cdna macroarray.
Clin Exp Metastasis
2002
.
19
:
391
399
.
Kim
,
S. K.
,
M.
Hebrok
, and
D. A.
Melton
.
Notochord to endoderm signaling is required for pancreas development.
Development
1997
.
124
:
4243
4252
.
Hebrok
,
M.
,
S. K.
Kim
, and
D. A.
Melton
.
Notochord repression of endodermal sonic hedgehog permits pancreas development.
Genes Dev
1998
.
12
:
1705
1713
.
Riobo
,
N. A.
,
K.
Lu
, and
C. P. Jr
Emerson
.
Hedgehog signal transduction: signal integration and cross talk in development and cancer.
Cell Cycle
2006
.
5
:
1612
1615
.
Beachy
,
P. A.
,
S. S.
Karhadkar
, and
D. M.
Berman
.
Tissue repair and stem cell renewal in carcinogenesis.
Nature
2004
.
432
:
324
331
.
Berman
,
D. M.
,
S. S.
Karhadkar
, and
A.
Maitra
.
et al
.
Widespread requirement for hedgehog ligand stimulation in growth of digestive tract tumours.
Nature
2003
.
425
:
846
851
.
Thayer
,
S. P.
,
M. P.
di Magliano
, and
P. W.
Heiser
.
et al
.
Hedgehog is an early and late mediator of pancreatic cancer tumorigenesis.
Nature
2003
.
425
:
851
856
.
Feldmann
,
G.
,
S.
Dhara
, and
V.
Fendrich
.
et al
.
Blockade of hedgehog signaling inhibits pancreatic cancer invasion and metastases: a new paradigm for combination therapy in solid cancers.
Cancer Res
2007
.
67
:
2187
2196
.

The authors have no relevant financial interest in the products or companies described in this article.

Author notes

Reprints: Christine A. Iacobuzio-Donahue, MD, PhD, The Johns Hopkins Medical Institutions, Department of Pathology, Division of Gastrointestinal/Liver Pathology, 1550 Orleans St, CRB2, Room 343, Baltimore, MD 21231 (ciacobu@jhmi.edu)