Invasive Pancreatic Adenocarcinoma Research Articles

Macrophage inflammatory protein-3α promotes pancreatic cancer cell invasion

Human CC chemokine Macrophage Inflammatory Protein-3alpha (MIP-3alpha) directs inflammatory cell migration through its binding to the transmembrane receptor CCR6. MIP-3alpha has recently been shown to promote tumor cell migration in pancreatic adenocarcinoma by up-regulation of matrix metalloproteinases (MMPs). We hypothesized that MIP-3alpha promotes pancreatic cancer invasion through the up-regulation of MMP-9, a Type 4 collagenase. Immunohistochemistry and RT-PCR confirmed the presence of MIP-3alpha in PANC-1 cells, a human pancreatic adenocarcinoma cell line. MIP-3alpha stimulated the production of both latent and active forms of MMP-9 in PANC-1 by Western analysis. Tumor cell invasion was then evaluated using a modified Boyden chamber invasion assay. MIP-3alpha promoted a dose-dependent increase in pancreatic cancer cell invasion (P < 0.05) at 100 ng/ml. The activity at the putative MIP-3alpha receptor, CCR6, was demonstrated by receptor blockade. Anti-CCR6 antibody and anti-MMP-9 antibody inhibited MIP-3alpha-stimulated PANC-1 cell invasion of collagen to 37% and 35% of control, respectively (P < 0.05). MIP-3alpha, through its CCR6 receptor, promotes tumor cell invasion by the up-regulation of MMP-9. Molecular based therapy aimed at the inhibition of MIP-3alpha activity through the CCR6 receptor may serve as a future target to prevent tumor cell invasion in pancreatic adenocarcinoma.

E-cadherin and β-catenin expression in pancreatic intraepithelial neoplasia and pancreatic adenocarcinoma

Objective: To investigate the significance of abnormal E-cadherin and β-catenin expression in pancreatic intraepithelial neoplasia (PanIN) and pancreatic adenocarcinoma. Methods:Pancreatic samples of 156 cases were retrospectively studied from surgery and autopsy in Changhai hospital from January 2001 to December 2003, from which tissue microarray blocks containing 129 PanIN-1A lesions, 104 PanIN-1B lesions, 22 PanIN-2 lesions, 11 PanIN-3 lesions, and 121 pancreatic ductal adenocarcinomas and corresponding paracancerous tissues were constructed. EnVision method of immunohistochemistry was used to detect the E-cadherin and β-catenin expression. The correlation between the abnormal E-cadherin and β-catenin expression and clinicopathological parameters was analysed. Results: The rate of E-cadherin abnormal expression was significant in ductal adenocarcinomas compared with the PanIN lesions and normal ducts(64.5%,32.3%,0%), moreover, the rate of E-cadherin abnormal expression was in relation to differentiation, lymph node metastasis and perineural invasion of pancreatic adenocarcinoma(P0.05). There was remarkable increase in the E-cadherin cytoplasmic expression in PanIN lesions and ductal adenocarcinomas compared with normal ducts. The rate of β-catenin abnormal expression was found to be related with lymph node metastasis and perineural invasion of pancreatic adenocarcinoma(P0.05). The expression of β-catenin cytoplasm and/or nucleus was significant in high-grade PanIN lesions and ductal adenocarcinomas compared with low grade PanIN lesions or normal ducts(P0.05). There was a positive relationship between the E-cadherin and β-catenin expression in PanIN lesions and ductal adenocarcinomas (P0.01, P0.05). Conclusion: There is aberration in the expression of the E-cadherin and β-catenin in PanIN lesions and ductal adenocarcinomas, suggesting the E-cadherin and β-catenin changes is not only related with the biological action and prognosis, but also involved in pancreatic carcinogenesis.

Chronic exposure of transforming growth factor beta 1 confers a more aggressive tumor phenotype through downregulation of p21WAF1/CIP1 in conditionally immortalized pancreatic epithelial cells

BackgroundRecent studies have demonstrated that transforming growth factor beta 1 (TGF-β1) expression is markedly enhanced in invasive ductal pancreatic adenocarcinomas, although the precise role of TGF-β1 in pancreatic carcinogenesis remains unclear. We analyzed TGF-β1 expression in pancreatic intraepithelial neoplasias (PanINs) and the effects of chronic TGF-β1 exposure on conditionally immortalized pancreatic epithelial (IMPE) cells. MethodsSixty-one PanIN lesions were immunohistochemically stained with a polyclonal rabbit antibody against human TGF-β1. Growth-inhibitory effects of short-term exposure to TGF-β1 were examined in IMPE cells. IMPE cells resistant to TGF-β1 (IMPE-Tr cells) were generated by continuous exposure to 1 ng/mL of TGF-β1 for more than 50 days. Phenotypic alterations of IMPE-Tr cells were examined by soft agar and Matrigel assay and Western blot analysis. IMPE and IMPE-Tr cells were injected subcutaneously into nude mice for an in vivo tumorigenicity assay. ResultsForty-six percent of PanINs (28/61) were positive for TGF-β1 expression, whereas all the epithelia of normal pancreatic ducts were negative. TGF-β1 treatment showed the marked growth-inhibitory effects (>75%) in IMPE cells, whereas its effects were not observed in IMPE-Tr cells. IMPE-Tr cells were more spindle shaped compared with IMPE cells. In soft agar and Matrigel, formations of many colonies were observed in IMPE-Tr cells, but not in IMPE cells. Interestingly, the expression of p21WAF1/CIP1 was induced by short-term exposure to TGF-β1 in IMPE cells, whereas the induction was decreased in IMPE-Tr cells. All of the IMPE-Tr cell-injected mice (5/5) had subcutaneous tumors, although no tumor was found in the IMPE cell-injected mice. ConclusionsTGF-β1 expression in PanINs and neoplastic transformation of IMPE cells by long-term exposure to TGF-β1 suggest that TGF-β1 may act as a tumor promoter in the early stage of pancreatic carcinogenesis.

Inflammation and Cancer V. Chronic pancreatitis and pancreatic cancer.

Pancreatic inflammation appears to increase the risk of pancreatic cancer. This observation is striking in the hereditary pancreatitis kindreds but also occurs in alcoholic, idiopathic, and tropical chronic pancreatitis and cystic fibrosis. However, the mutations associated with hereditary pancreatitis or cystic fibrosis are not found in sporadic pancreatic adenocarcinomas, suggesting that the effects are indirect by causing recurrent pancreatitis and chronic inflammation. The process of mutation accumulation and clonal expansion that is required for development of invasive pancreatic adenocarcinoma must therefore be accelerated in chronic pancreatitis to account for the high incidence of pancreatic cancer in these patients.

Macrophage inflammatory protein-3alpha (mip-3alpha) promotes pancreatic cancer cell invasion by upregulation of matrix metalloproteinase-9 (MMP-9) production

Introduction: MIP-3α has been shown to promote tumor cell migration by upregulation of MMPs. We hypothesize that MIP-3α, by binding to the transmembrane receptor CCR6, promotes pancreatic cancer invasion through the upregulation of MMP-9, a type IV collagenase that plays a crucial role in tumor progression. Methods: RT-PCR for MIP-3α was performed on PANC-1, human pancreatic adenocarcinoma cells. The effect of MIP-3α on MMP-9 production by PANC-1 cells was evaluated by Western analysis. Tumor cell invasion was evaluated using a modified Boyden chamber invasion assay. Results: RT-PCR confirmed the presence of MIP-3α in PANC-1 cells. MIP-3α stimulated the production of both latent and active forms of MMP-9 as demonstrated by Western analysis (see Figure). MIP-3α promoted a dose-dependent increase in pancreatic cancer cell invasion (see Table). TABLE—ABSTRACT 36Summation of Invasive Cells in Five[MIP-3α] ng/ml01050100Invasive cells9.0 ± 6.113.0 ± 2.616.7 ± 4.523.3 ± 5.0∗ Anti-CCR6 Ab and anti-MMP-9 Ab inhibited MIP-3α stimulated PANC-1 invasion of collagen to 37% and 35% of control, respectively (p < 0.05). Conclusion: MIP-3α, through its CCR6 receptor, causes a dose-dependent increase in tumor cell invasion by the upregulation of MMP-9. Molecular based therapy aimed at the inhibition of MIP-3α through the CCR6 receptor may serve as a future target to prevent tumor cell invasion in pancreatic adenocarcinoma.

Multicomponent Analysis of the Pancreatic Adenocarcinoma Progression Model Using a Pancreatic Intraepithelial Neoplasia Tissue Microarray

A multistep model for pancreatic adenocarcinoma has been proposed recently. In this model, well-defined, noninvasive ductal lesions are recognized as precursors of invasive cancer and have been classified under the nomenclature of pancreatic intraepithelial neoplasia, or PanIN. Increasing evidence suggests that PanINs represent true neoplasms of the pancreatic ductal epithelium, accumulating histologic and genetic abnormalities in their progression toward invasive cancer. We have constructed a tissue microarray containing 55 PanIN lesions of all histologic grades in order to perform a multicomponent analysis of the pancreatic adenocarcinoma progression model. The protein products of 14 genes encompassing a variety of functional classes, such as tumor suppressor genes (p53, Smad4/Dpc4), oncogenes (β-catenin), cell cycle antigens (p16, cyclin D1), proliferation antigens (Ki-67, topoisomerase II alpha), and epithelial apomucins (MUC1, MUC2, MUC5), as well as “novel” genes described as differentially up-regulated in invasive pancreas cancer by global microarray expression analysis (mesothelin, prostate stem cell antigen, fascin, and 14-3-3σ), were analyzed by immunohistochemistry on the PanIN tissue microarray. Comparison of the results from the current study with previously published data performed on routine histologic sections of PanINs demonstrates that tissue microarrays are a valid platform for molecular analysis not only of invasive cancers but of precursor lesions as well. In addition, this study demonstrates that molecular abnormalities in PanINs are not random but can usually be stratified into “early” changes (e.g., expression of MUC5 and prostate stem antigen, or loss of p16), “intermediate” changes (e.g., expression of cyclin D1), and “late” changes (e.g., expression of p53, proliferation antigens, MUC1, mesothelin, and 14-3-3σ, or loss of Smad4/Dpc4). Understanding the molecular pathogenesis of precursor lesions of invasive pancreatic adenocarcinomas using a high-throughput tissue microarray–based approach is a valuable adjunct to designing rational strategies for early detection of this lethal neoplasm.

Telomere Shortening Is Nearly Universal in Pancreatic Intraepithelial Neoplasia

A multistep model of carcinogenesis has recently been proposed for pancreatic ductal adenocarcinomas. In this model, noninvasive precursor lesions in the pancreatic ductules accumulate genetic alterations in cancer-associated genes eventually leading to the development of an invasive cancer. The nomenclature for these precursor lesions has been standardized as pancreatic intraepithelial neoplasia or PanIN. Despite the substantial advances made in understanding the biology of invasive pancreatic adenocarcinomas, little is known about the initiating genetic events in the pancreatic ductal epithelium that facilitates its progression to cancer. Telomeres are distinctive structures at the ends of chromosomes that protect against chromosomal breakage-fusion-bridge cycles in dividing cells. Critically shortened telomeres can cause chromosomal instability, a sine qua non of most human epithelial cancers. Although evidence for telomeric dysfunction has been demonstrated in invasive pancreatic cancer, the onset of this phenomenon has not been elucidated in the context of noninvasive precursor lesions. We used a recently described in situ hybridization technique in archival samples (Meeker AK, Gage WR, Hicks JL, Simon I, Coffman JR, Platz EA, March GE, De Marzo AM: Telomere length assessment in human archival tissues: combined telomere fluorescence in situ hybridization and immunostaining. American Journal of Pathology 2002, 160:1259-1268) for assessment of telomere length in tissue microarrays containing a variety of noninvasive pancreatic ductal lesions. These included 82 PanIN lesions of all histological grades (24 PanIN-1A, 23 PanIN-1B, 24 PanIN-2, and 11 PanIN-3) that were selected from pancreatectomy specimens for either adenocarcinoma or chronic pancreatitis. Telomere fluorescence intensities in PanIN lesions were compared with adjacent normal pancreatic ductal epithelium and acini (62 of 82 lesions, 76%), or with stromal fibroblasts and islets of Langerhans (20 of 82 lesions, 24%). Telomere signals were strikingly reduced in 79 (96%) of 82 PanINs compared to adjacent normal structures. Notably, even PanIN-1A, the earliest putative precursor lesion, demonstrated a dramatic reduction of telomere fluorescence intensity in 21 (91%) of 23 foci examined. In chronic pancreatitis, reduction of telomere signal was observed in all PanIN lesions, whereas atrophic and inflammatory ductal lesions retained normal telomere length. Telomere fluorescence intensity in PanIN lesions did not correlate with proliferation measured by quantitative Ki-67-labeling index or topoisomerase IIalpha expression. Thus, telomere shortening is by far the most common early genetic abnormality recognized to date in the progression model of pancreatic adenocarcinomas. Telomeres may be an essential gatekeeper for maintaining chromosomal integrity, and thus, normal cellular physiology in pancreatic ductal epithelium. A critical shortening of telomere length in PanINs may predispose these noninvasive ductal lesions to accumulate progressive chromosomal abnormalities and to develop toward the stage of invasive carcinoma.

MUC4 expression increases progressively in pancreatic intraepithelial neoplasia.

Pancreatic adenocarcinoma is believed to develop from histologically identifiable intraductal lesions known as pancreatic intraepithelial neoplasias (PanINs) that undergo a series of architectural, cytologic, and genetic changes, a progression model similar to the adenoma-carcinoma sequence in the colon. The apomucin MUC4 has been implicated in invasive pancreatic adenocarcinoma. MUC4 expression is not detectable at the RNA level in normal pancreas but is detectable at high levels in invasive pancreatic adenocarcinoma. We documented the pattern of expression of MUC4 in PanINs by studying a series of 71 PanIN lesions immunohistochemically using a new monoclonal antibody to MUC4. Five (17%) of 30 PanIN-1 lesions, 10 (36%) of 28 PanIN-2 lesions, 11 (85%) of 13 PanIN-3 lesions, and 25 (89%) of 28 invasive adenocarcinomas labeled with the MUC4 antibody used in the study. In addition, afew nonneoplastic lesions labeled with the MUC4 antibody, including reactive ducts in chronic pancreatitis, atrophic ducts filled with inspissated secretions, and ducts showing squamous metaplasia. Our data help establish the patterns of MUC4 expression in neoplastic precursors in the pancreas and add further support to the progression model for pancreatic adenocarcinoma.

Neoplasms of the Ampulla of Vater with Concurrent Pancreatic Intraductal Neoplasia: A Histological and Molecular Study

Adenoma and adenocarcinoma of the ampulla of Vater are uncommon neoplasms of the gastrointestinal tract. Only one report has analyzed the relationship between ampullary adenocarcinoma and pancreatic intraductal neoplasia (PanIN), the precursor lesion of pancreatic adenocarcinoma. An association between PanIN and ampullary adenoma has not been reported previously. Case reports have documented the progression of PanIN to invasive pancreatic adenocarcinoma. We reviewed five resected ampullary adenoma and 17 ampullary adenocarcinoma cases and evaluated the pancreas for PanIN. Pancreatic sections from 35 autopsies were reviewed as a control group. Immunohistochemistry for overexpression of p53 and COX-2 proteins was performed in selected cases, as was PCR analysis for K-ras mutations. Follow-up clinical data were obtained. All 22 ampullary neoplasms were associated with PanIN, which was high grade in two (40%) adenoma cases and seven (41%) adenocarcinoma cases. In 16 (73%) evaluable cases, PanIN extended to the pancreatic resection margin; two of which had high grade PanIN. Among the autopsy controls eight (23%) had low-grade PanIN. Seven of the 22 ampullary cases but none of the autopsy controls had coexistent pancreatitis. A smoking history was present in two of four autopsy cases in which this history was available. Overexpression of the p53 and COX-2 proteins was present in only one case of high-grade PanIN. K-ras mutations were present in four of four of the PanIN lesions evaluated, including one autopsy case. Clinical follow-up revealed no progression of PanIN to invasive carcinoma in the remnant pancreas, although the follow-up period was too short to adequately assess that risk (an average of 3.8 y for adenoma cases and 2.5 y for adenocarcinoma cases). We conclude that adenomas and carcinomas of the ampulla are associated with PanIN, and often high-grade PanIN. Although its malignant potential has not been fully established, PanIN is underreported and often unrecognized. PanIN may be analogous to colorectal adenoma in that both are prevalent in the older adult population, but few progress to carcinoma.

Overexpression of midkine in pancreatic duct adenocarcinomas induced by N-Nitrosobis(2-oxopropyl)amine in hamsters and their cell lines.

The expression of midkine (MK) was investigated in pancreatic ductal hyperplasias, atypical hyperplasias and adenocarcinomas induced by N‐nitrosobis(2‐oxopropyl)amine (BOP) in hamsters, and in hamster ductal adenocarcinoma cell lines (HPD‐1NR, ‐2NR and ‐3NR). MK mRNA was clearly overexpressed in invasive pancreatic duct adenocarcinomas (PCs) and the three cell lines as assessed by northern blot analysis, and MK protein expression increased from ductal hyperplasia through atypical hyperplasias, intraductal carcinomas and invasive PCs by immunohistochemistry. The extent of overexpression of MK mRNA in PCs was almost the same as in hamster whole embryonic tissue. MK is reported to be a retinoid‐responsive gene, but MK mRNA expression was not affected by treatment with all‐trans retinoic acid (tRA) or N‐(4‐hydroxyphenyl)retinamide (4‐ HPR) in HPD‐1NR cells. The results thus suggest that MK expression is involved in the development and progression of pancreatic ductal adenocarcinomas induced by BOP in hamsters, with loss of upregulation by retinoic acid.

Immunohistochemical evaluation of K- ras, p53, and HER-2/ neu expression in hyperplastic, dysplastic, and carcinomatous lesions of the pancreas: Evidence for multistep carcinogenesis

The pathobiology of precursor lesions leading to invasive pancreatic adenocarcinoma remains a controversial area, but knowledge of the mechanisms of tumorigenesis may lead to possibly earlier detection, prevention, and treatment in the future. We hypothesize that ductal hyperplasia and dysplasia of the pancreas represent precursor lesions and are part of a continuous developmental spectrum evolving into ductal adenocarcinoma of the pancreas. To further define this sequence, we studied the immunohistochemical markers HER-2/ neu, K- ras, and p53 in 15 adenocarcinomas and 15 nonmalignant specimens of the pancreas. The 15 nonmalignant specimens of the pancreas included both normal pancreas and chronic pancreatitis. Overall, HER-2/ neu was positive in normal ducts, ductal hyperplasia, dysplasia, and carcinoma cells in 0 of 30, 11 of 20 (55%), 10 of 15 (67%), and 12 of 15 (80%), respectively, with progressive increase in the intensity of staining; p53 was positive in 1 of 30 (3%), 0 of 20, 3 of 15 (20%), and 13 of 15 (80%), respectively, and K- ras was positive in 1 of 30 (3%), 6 of 20 (30%), 11 of 15 (73%), and 8 of 15% (53%), respectively. These data support the hypothesis that ductal hyperplasia and dysplasia of the pancreas represent precursor lesions, and, in a fashion similar to that in colorectal tumorigenesis, pancreatic cancer seems to accumulate progressive genetic alterations.

Decreased membrane distribution of e-cadherin results in poor cell-cell adhesion in a highly invasive pancreatic adenocarcinoma, PC-1

Decreased membrane distribution of e-cadherin results in poor cell-cell adhesion in a highly invasive pancreatic adenocarcinoma, PC-1

Pictorial Essay. Dual-Phase Helical CT of Locally Invasive Pancreatic Adenocarcinoma

Dual-phase helical CT permits imaging of the pancreas and the peripancreatic structures in the arterial dominant and portal venous phases of enhancement, providing information crucial in the assessment of the local extent of pancreatic adenocarcinoma. This essay reviews the dual-phase helical CT findings of local extension that preclude potentially curative surgery, including vascular involvement, ligamentous or mesenteric invasion, extension of the tumor to involve the stomach or duodenum, and invasion of adjacent solid organs.

The effect of the somatostatin analogue SMS 201-995 on experimental pancreatic carcinogenesis in the Syrian golden hamster

Somatostatin analogues have been suggested as possible therapy for human pancreatic cancer. This paper investigates the effect of the somatostatin analogue SMS 201-995 (Sandoz) in the Syrian golden hamster model of nitrosamine-induced pancreatic carcinogenesis. Step-wise increasing doses of i.v. SMS 201-995 suppressed pancreatic juice output from a median basal value of 212 mg/kg body wt/h (Q1:Q3 = 121:334) to a median basal value of 70 mg/kg body wt/h during infusion of 5 micrograms/kg body wt/h of SMS 201-995 (Q1:Q3 = 64:102, P less than 0.05). Chronic s.c. injection of 5 and 10 micrograms/kg body wt SMS 201-995 twice daily for 3 days each week, did not affect pancreatic wet wt or pancreatic total DNA content after 1 or 6 weeks of treatment when compared to controls. The most interesting and unexpected finding in our study was that SMS 201-995 seemed to promote pancreatic carcinogenesis when administered in low dosage. More SMS 201-995 treated animals receiving 5 micrograms/kg body wt developed invasive pancreatic adenocarcinoma than controls after 15 weeks of carcinogen (4/10 animals versus 0/10, P less than 0.05, Fisher's exact test) and pancreatic involvement by tumour was more extensive (17/75 pancreatic blocks affected versus 0/71, P less than 0.001). When carcinoma in situ and microcarcinomata were analysed with invasive lesions, animals injected with 5 micrograms/kg body wt SMS 201-995 were still significantly more affected than controls (33/75 blocks versus 9/71, P less than 0.001). Hamsters injected with the higher SMS 201-995 dose (10 micrograms/kg body wt) did not show any increase in malignancy over the controls. These results suggest that the effect of SMS 201-995 on pancreatic carcinogenesis in the Syrian hamster is complex and varies with dose administered. Further work is required before its use on man can be justified.

Streptozotocin prevents development of nitrosamine-induced pancreatic cancer in the Syrian hamster.

Administration of the nitrosamine carcinogen N-nitroso-bis (2-oxopropyl) amine (BOP) by subcutaneous injection (5 mg/kg/week) led to the development of invasive pancreatic ductular adenocarcinoma in 100% of normal Syrian hamsters by 24 weeks. Pretreatment of a second group of hamsters with the beta-cell toxin streptozotocin in a diabetogenic dose (50 mg/kg i.p. X 3) completely prevented the development of pancreatic cancer when BOP was subsequently administered. The mechanism of blockade by streptozotocin is unknown. This study suggests the potential importance of the endocrine pancreas in exocrine pancreatic carcinogenesis.