anaplastic cancers parallels the advanced loss of cellu- lar differentiation in cultured islets, where the cells lose every known islet-cell marker. Clinical studies supporting the role of islets in pancreatic carcinogenesis Although it has been known for almost a century that nearly 80% of patients with pancreatic cancer have im- paired glucose metabolism, either frank diabetes or impaired glucose tolerance (IGT), the reason has re- mained a mystery. Remarkably, the degree of IGT and diabetes in these patients has been known to vary. Some patients require insulin treatment whereas others do not. Fasting serum glucose levels may be in the normal range, but an oral glucose tolerance test may yield a di- agnosis of IGT. On the other hand, a small subset of pa- tients shows normal glucose metabolism. It is possible that differences in the patient population are the reason that morphologic and molecular biological approaches have not provided clues for understanding the biology of this dismal disease. The association between diabetes and pancreatic cancer has remained a matter of controversy. Accord- ing to recent studies, IGT or diabetes mellitus develops shortly before the clinical manifestations of the disease or is diagnosed at the first clinical admission. There are, however, a few who believe that diabetes is a predispos- ing factor, especially in cases where diabetes is present for more than 5 years before the diagnosis of cancer. Since the latency of pancreatic cancer is unclear, and the development of some cancers seems to take as long as 10 years, the role of diabetes as a predisposing factor remains questionable. Consequently, it appears that the development of pancreatic cancer is associated with the abnormality in islet cell function. Some suggested mechanisms include the primary alteration of islet cells by the carcinogen or secondary damage by cancer cells, either directly or via the production of substances that affect islet cell function. Experimental studies in the hamster model described above and anecdotal observations indicate that islet cells may also play a role in pancreatic carcinogenesis in humans. This is highlighted by the development of altered glucose metabolism in small tumors that are located in the periphery of the pancreas but which do not cause chronic pancreatitis and in the very early stages of cancer development. The involvement of islet cells in pancreatic carcinogenesis explains at least some of the clinical observations. Based on the published data and our experience, glucose intolerance is a pancreatic cancer-associated symptom as well as the result of the primary alteration of islet-cell function and differentiation in response to causative carcinogens. Experimental studies have shown that glucose intolerance coincides with the first appearance of microscopic pancreatic tumors. Studies have also shown that changes in islet hormones accom- PART III 370 Figure 44.5 An enlarged islet in a patient with pancreatic cancer. Malignant glands have replaced most islet cells without any signs of islet cell destruction, including invasion of the islet cells and the surrounding tissue. In some areas intact islet cells are present within the malignant epithelium (e.g., lower right area). Anti-insulin antibody, ABC (avidin–biotin complex) method, ¥ 120. Figure 44.6 Papillary projection of a well-differentiated pancreatic adenocarcinoma. Numerous insulin-producing cells are seen at the base and within the papillary fold. Anti- insulin, ABC (avidin–biotin complex) method, ¥ 120. pany the early development of pancreatic cancers. These hormone changes and insulin resistance resem- ble the metabolic changes in pancreatic cancer patients. In humans, IGT or diabetes has been noticed in small, localized, and early pancreatic cancer. In Japanese patients, IGT was the only abnormality in patients with small pancreatic cancer. The overwhelming opinion that pancreatic cancer develops from ductal epithelium might be the reason why islet-cell alterations in pancreatic cancer patients have not been the focus of research. When we studied the pattern of islet cells immunohistochemically in 14 pancreatic cancer specimens, 14 chronic pancreatitis samples, and 10 normal pancreata as controls, we found that 10 of 14 cancer specimens showed a signifi- cant loss of islet b cells. Of the 10 cases, IGT was con- firmed in four but no information was available about glucose metabolism in the remaining cases. The inci- dence of islet-cell alterations in our material (72%) correlates with the frequency of abnormal glucose metabolism in pancreatic cancer patients. Remarkably, most altered islet cells were in the vicinity of the cancer. In only one case was the abnormality also found in an area remote from the cancer. Since tumor-free pan- creatic tissues were available in only five cases, the frequency of islet cell alteration in the teletumoral area could not be determined. Other noteworthy findings associated with this abnormality were the signs of al- tered islet cell differentiation, including the formation of intrainsular ductular structures and the expression of tumor-associated antigens CA-19-9, TAG-72, and/ or DU-PAN-2 in islet cells and intrainsular ductular cells (see Fig. 44.3). This finding indicates that in these patients islet cells have the ability to form an abnormal cell population. Possible mechanism of altered glucose metabolism in pancreatic cancer It has been proposed that amylin, a peptide with a molecular weight of 2030, or other yet unknown sub- stances released from cancer cells are responsible for the development of IGT. Because we believe that most cancers arise from altered islet cells, the production of these substances from cancer cells is self-explanatory. Cancer cells are known to inherit some of the biological properties of the cells from which they are derived. In- deed, several studies show the expression of neuroen- docrine markers in pancreatic cancer cells. From a pathophysiologic point of view, the production of dia- betogenic material from islet cells appears more plausi- ble, as it is well known that islet cells have the potential to produce many different pancreatic and extrapancre- atic peptides simultaneously. They also have the ability to shift from synthesis of one hormone to synthesis of another. A good example is the coproduction and core- lease of insulin and amylin, the synchrony of which is altered in pancreatic cancer. The improvement in IGT and diabetes after tumor resection (70% pancreatecto- my or curative resection) by no means indicates that it was the tumor that produced the diabetogenic sub- stances, because removal of cancer tissue also removes the altered islet cells that may actually have produced the diabetogenic material. Moreover, we must be aware that nearly all well-differentiated pancreatic cancers contain endocrine cells, sometimes in remarkably high numbers (Fig. 44.7), which could also be the source of altered hormone production and which are also re- moved with the cancer. For example, although tumor extracts from diabetic patients with pancreatic cancer showed a marked reduction of glycogen synthesis in skeletal muscles, examination of the tumor revealed that tumor tissue contained islet hormones. Although from a clinical standpoint the issue of whether the diabetogenic material is produced by cancer cells or al- tered islet cells is trivial, elucidation of the mechanism is CHAPTER 44 371 (a) (b) Figure 44.7 Presence of a large number of islet cells within the malignant epithelium. (a) Many b cells are incorporated within the glandular structures. Anti-insulin antibody, ABC method, ¥ 25. (b) Malignant glandular structures containing more endocrine than cancer cells. Multilabeling technique, ¥ 120. crucial to understanding the biology of the disease and in planning future diagnostic and therapeutic modalities. Differences in the clinical expression of pancreatic cancer Although it appears that alteration of glucose metabo- lism can provide a diagnostic marker, some observa- tions complicate the issue. According to clinical observations, only 60–70% of patients develop IGT or diabetes and the minority (30–40%) do not. Although IGT improves after surgery in many patients, in some it does not or it gets even worse. There are conflicting reports and inadequate information on the incidence of peripheral insulin resistance, IGT, and diabetes before and after surgery. According to one study, 59% of pancreatic cancer patients with either diabetes (45%) or IGT (14%) show improvement after curative surgery, whereas studies by Permert et al., using a hyperglycemic clamp method, show normalization of IGT and improvement of diabetes in around 60% of patients. Consequently, it can be as- sumed that 10–40% of pancreatic cancer patients ei- ther do not show any improvement of the abnormality after surgery or IGT becomes worse. The latter figures could be even higher if one considers that postoperative improvement of IGT and diabetes could be due to the postoperative physical condition and dietary regimens of the patients rather than the consequence of tumor re- moval. It is unclear whether the observed improvement is just temporary or if the abnormality reappears at the time of tumor recurrence. Although many reasons could be responsible for the lack of postoperative improvement of glucose metabolism in the subset of patients, it is highly possible that altered islet cells producing diabetogenic substances exist in a teletu- moral area not removed by surgery or some hidden (metastatic) tumors are left behind, for example in the liver. Since in a follow-up study glucose homeostasis in- creasingly worsened in patients who did not have cura- tive surgery, the extent of the tumor and/or altered islets seems to be responsible for glucose metabolism. There are, as yet, no studies examining the extent of islet cell alteration within, around, and remote from cancer. Also, there are limited follow-up studies of patients after surgery. Possible etiologic factors for islet-cell alteration in pancreatic cancer The results of our 30 years of experience in human and experimental pancreatic cancer has led us to believe that islet cells are the primary targets of carcinogens. In our view, all pancreatic tumors, endocrine or exocrine, are derived from islets. The structure of the carcinogen determines the phenotypic expression of the ensuing tumors. Streptozotocin, a nitrosamide, produces islet-cell tumors, whereas BOP, a nitrosamine, induces a ductal type of tumor. In hamsters and humans, cultured islet cells transdifferentiate into ductal cells. In hamsters, BOP treatment of isolated purified islets leads to tumor cells that grow in vivo as ductal adeno- carcinoma. When we treat cultured human ductal and islet cells with BOP, only the treated islet cells are able to grow in a serum-free medium and show K-ras mutation, a marker for pancreatic cancer (unpublished results). In an ongoing study we are following the characteristics of these cells and expect their malignant transformation. The most convincing support for our view is the finding that all drug-metabolizing enzymes, which are believed to be involved in the metabolism of environmental carcinogens, including tobacco-specific carcinogen, nitrosamines, polycyclic aromatic com- pounds, and aromatic amines, are primarily or exclusively expressed in the islet cells of humans and laboratory species. Considering the anatomy of the blood supply of the pancreas, where a major portion of the arterial blood goes to islets before nourishing the exocrine pancreas, the presence of drug-metabolizing enzymes in islet cells is understandable. Hence, islet cells seem to play the role of pancreatic filters. The availability of these enzymes makes islet cells the pri- mary target of blood-borne carcinogens. Because most of these enzymes are present in a higher concentration or exclusively in islet cells in the head of the pancreas, the frequent occurrence of pancreatic cancer in the head may be explained. Carcinogen-induced alter- ations in the islets in teletumoral regions of the pan- creas could be the reason for the altered production of hormones and, hence, the maintenance of IGT after tumor removal. This explanation, however, is not conclusive because not all pancreatic cancer patients develop a glucose metabolic abnormality. Is this related to the different biology of cancer, as has been suggested by a study where a correlation was found between the PART III 372 degree of IGT severity and the histologic type of can- cer? Is this because tumors develop from islets in pa- tients with IGT or diabetes and, in a minority of the patients, from other cells? Or could this be related to the severity and extent of islet-cell damage? Neverthe- less, the data suggest that, with regard to the glucose metabolic alteration, there are at least three subsets of pancreatic cancer patients, possibly with tumors of different biology. The published data and our own experience suggest the following subsets (Fig. 44.4): 1 pancreatic cancer patients without IGT or diabetes (IGT–, about 20–30%); 2 pancreatic cancer patients with IGT or diabetes (IGT+, about 70–80%), whose glucose intolerance or diabetes improves postoperatively (IGT+/–); 3 patients in whom the abnormality does not or only slightly improves (IGT+/+) after tumor resection. Possible mechanism of differing clinical presentation of pancreatic cancer Reasons for the glucose metabolic abnormality in pan- creatic cancer are not well understood. The suggestion that islet-cell destruction by cancer cells is the principal cause has been refuted, mainly because even small and localized tumors in the head of the pancreas are associ- ated with abnormal glucose tolerance. A few studies dealing with the alteration of islet hor- mones at the tissue level have found a reduction in the number of b cells in pancreatic cancer patients. No information is available on the frequency and extent of the process, and its specificity for pancreatic cancer. The question of specificity is important because about 45–75% of patients with chronic pancreatitis also develop abnormal glucose tolerance or frank diabetes mellitus. Consequently, it is reasonable to assume that damage to the islets by scar tissue and inflammation, which are also associated with pancreatic cancer, could be the underlying mechanism. Clearly, disturbance of the subtle balance between exocrine and endocrine tissue by cancer, with asso- ciated inflammation and sclerosis, is expected to lead to deregulation of hormone secretion. However, because even localized and small tumors in the head of the pan- creas not affecting islet-rich areas of the organ cause the glucose metabolic abnormality, it is likely that a factor or factors produced by cancer cells play a role. This view is supported by the finding that surgical removal of tumor by 85–90% pancreatectomy improves dia- betes and normalizes glucose metabolism. Several clinical studies have shown significant changes in the serum levels of islet hormones in pancre- atic cancer patients, but little is known about the pat- terns of islets at the tissue level. In one study a reduction in b cells has been reported but the extent of the alter- ations and their specificity for pancreatic cancer have not been investigated. The latter issue deserves par- ticular attention, because glucose-abnormality and diabetes also occurs in chronic pancreatitis. Therefore, we systematically examined the patterns of islets in pancreatic cancer in comparison with chronic pancre- atitis and the normal pancreas. We selected archival pancreatic cancer specimens that had tumor-free areas close to and remote from the cancer because, as stated earlier, it is believed that factors released by cancer cells affect the islets directly via a paracrine pathway. In 10 pancreatic cancer specimens, a significant reduction in b cells was found. Also, in eight of them, a significant increase in a cells was found as well. This re- sult thus correlates with the incidence of pathologic serologic hormone levels in pancreatic cancer patients. Reasons for the lack of similar alterations in the four other patients are obscure. We could not find any corre- lation between islet alterations, sex, age, smoking habit, alcohol consumption, stage of the disease, and tumor morphology. We also did not find any significant changes in the islet-cell distribution in the chronic pan- creatitis specimens, even within sclerotic and fibrotic tissue. Therefore, the suggestion that fibrosis or sclero- sis associated with pancreatic cancer may have caused the b-cell loss by obstructing blood vessels could be excluded. Consequently, the described islet alteration appears to be specific for pancreatic cancer. Because pathologic islet hormone serum levels also occur in chronic pancreatitis patients, it seems that the mecha- nism of altered glucose metabolism in the two diseases differs. In chronic pancreatitis the abnormality seems to be due to altered insulin secretion, whereas in pan- creatic cancer the defect appears to be in the machinery of insulin synthesis as evidenced by the reduced levels of insulin and C-peptide as well as of amylin, which is nor- mally costored and cosecreted with insulin. Endocrine cells were found in the malignant epithelium in nine of our ten cases with b-cell alteration. Similar findings have been reported in up to 80% of cases. Also, the presence of nesidioblastosis in four of our ten cases with decreased b-cell number could reflect a compen- CHAPTER 44 373 satory process against b-cell loss. The question of why b cells in pancreatic cancer are exclusively affected remains to be investigated. The increase in a cells, which was more pronounced in cancer tissue from diabetics than in tissue from dia- betics without cancer, coincides with the serologic findings. The abnormality also differs from hormonal changes in chronic pancreatitis, where serum concen- trations of glucagon have been found to be reduced or normal. Nevertheless, in our chronic pancreatitis sam- ples we could not detect any alteration in the number of glucagon cells, possibly because it is the secretion of glucagon that is affected not the number of a cells. Also, contrary to clinical observations of increased so- matostatin levels in pancreatic cancer patients, we could not find any significant changes in the number of somatostatin cells. Whether the source of increased serum somatostatin is derived from pancreatic or ex- trapancreatic somatostatin cells remains to be seen. The greater alteration of islets within or immediately around cancer supports the hypothesis that factors re- leased by cancer cells play a role in this process. Because alterations like hydropic swelling were also found in tissues remote from cancer, although in lesser degree, a humoral pathway also seems to exist. Examination of pancreatic tissue further away from the cancer would clarify this. If this is found to be the case, the identifica- tion of the causative factor(s) released from cancer cells could present an early pancreatic cancer marker, espe- cially in view of the findings that an abnormality in glu- cose metabolism also occurs in small localized cancers. One of the reasons for the differing results in the pub- lished data and pancreatic hormone levels in pancreat- ic cancer patients could be the inclusion of different subsets of patients with pancreatic cancer in these studies. It may be that the anatomic location of the tumor plays a role in these differences. Tumors in the head region obstructing the main pancreatic duct can cause severe (secondary) chronic pancreatitis and, hence, diabetes. However, there are differences in dia- betes induced by chronic pancreatitis and pancreatic cancer. For example, diabetes improves after a 70% pancreatectomy in pancreatic cancer but not after sur- gical intervention in chronic pancreatitis. According to our recent studies, the size and cell constitution of islets are significantly different between primary pancreatitis and pancreatitis caused by cancer. Contrary to the islets in pancreatic cancer patients, which are of normal size or enlarged, about 95% of the islets in primary chronic pancreatitis measure less than 100 mm in diameter. Moreover, tumors developing in the upper and dorsal half of the head of the pancreas do not affect the pan- creatic duct very much and hence are not accompanied by a significant chronic pancreatitis. Whether these tumors cause diabetes is unclear. Another argument against the role of secondary pancreatitis in the induc- tion of diabetes derives from the experience that even patients with small tumors in the periphery of the pan- creas not causing chronic pancreatitis show abnormal glucose tolerance. Another major shortcoming of past studies is the lack of adequate control groups. There is not a single study that correlates the morphologic findings of both cancer and islet cells with plasma hormone levels of the pa- tients. To our knowledge, there is only one limited study that compares the hormone levels in pancreatic cancer patients with that of healthy and noninsulin- requiring diabetic persons: a low level of plasma amylin was found in patients with type II diabetes and in those with pancreatic cancer and diabetes, and an increased level of amylin in pancreatic cancer patients without diabetes. Such correlative studies could provide impor- tant data for an understanding of the disease. For example, a low insulin level in a patient with IGT and altered islet cells could reflect impaired insulin release or synthesis in the altered islets. In fact, an inverse rela- tionship has been found between the number of insulin cells in islets and the fasting plasma glucose level, sug- gesting that the alteration of islet cells is the primary cause of the glucose abnormality in these patients. Other studies also point to the primary alterations of islet cells, including reduced insulin and C-peptide re- sponse after glucose load and an increase in proinsulin secretion. The Sproinsulin/SC-peptide ratio, which has been found to be increased in pancreatic cancer with IGT but decreased after tumor removal, further sub- stantiates the functional alteration of islet cells. From a therapeutic point of view, the identification of a different subpopulation of pancreatic cancer patients is important because these patients could respond differently to therapeutic modalities. For example, does the genetic constitution of the tumors from the different pancreatic cancer subpopulations differ? Can the pattern of IGT help to better distinguish between sporadic and familial pancreatic cancer? If IGT is due to the substances released from islet cells or cancer cells, then is it expected that IGT+/– patients become IGT+/+ after tumor recurrence? If that is the PART III 374 case, the recurrence of the abnormality, and possibly its severity, could have predictive value. The existing re- sults indicate that the occurrence of diabetes in pancre- atic cancer cannot be explained by a single mechanism. The increase in peripheral insulin resistance, suppres- sion of insulin secretion, impaired proinsulin con- version, altered fat and carbohydrate metabolism, presence of acute or chronic pancreatitis, medications for underlying disease, altered nutritional habits, weight loss, and many other factors seem to play im- portant roles in the development and course of pancre- atic cancer. Conclusion Past attempts to develop early diagnostic modalities have proven useless. The expression of tumor-associat- ed antigens may have value in monitoring the disease but are unable to detect the cancer in early developmen- tal stages. Despite the promising molecular biological approach, the method lacks specificity as the K-ras mu- tation is not specific for pancreatic cancer and can be found in patients with chronic pancreatitis as well as in individuals without pancreatic diseases. The most so- phisticated imaging techniques are still unable to detect tumors less than 5 cm with accuracy. The frequent asso- ciation between pancreatic cancer and IGT offers the most logical approach for detecting small tumors in most patients. This method could be applied readily in individuals prone to pancreatic cancer, including members of pancreatic cancer and hereditary chronic pancreatitis. Our studies pointing to the role of islets in pancreatic cancer and in the development of altered glucose metabolism should be further investigated. The development of a multidisciplinary program involving researchers in various fields of medicine, toxicology, nutrition, cellular and molecular biology, and epidemi- ology is a necessary step in revealing the true nature of this deadly disease. Recommended reading Ahren B, Andren-Sandberg A. Glucose tolerance and insulin secretion in experimental pancreatic cancer in the Syrian hamster. Res Exp Med 1993;193:21–26. Bonner-Weir S, Baxter LA, Schuppin GT, Smith FE. A second pathway for regeneration of adult exocrine and endocrine pancreas. A possible recapitulation of embryonic develop- ment. Diabetes 1993;42:1715–1720. Bouwens L. Transdifferentiation versus stem cell hypothesis for the regeneration of islet beta-cells in the pancreas. Microsc Res Tech 1998;43:332–336. Bouwens L, Kloppel G. Islet cell neogenesis in the pancreas. Virchows Arch 1996;427:553–560. Cersosimo E, Pisters PW, Pesola G, McDermott K, Bajorunas D, Brennan MF. Insulin secretion and action in patients with pancreatic cancer. Cancer 1991;67:486–493. Gittes GK, Galante PE, Hanahan D, Rutter WJ, Debase HT. Lineage-specific morphogenesis in the developing pancreas: role of mesenchymal factors. Development 1996;122:439– 447. Gullo L, Ancona D, Pezzilli R, Casadei R, Campione O. Glu- cose tolerance and insulin secretion in pancreatic cancer. Ital J Gastroenterol 1993;25:487–489. Jonsson J, Carlsson L, Edlund T, Edlund H. Insulin-promoter- factor 1 is required for pancreas development in mice. Nature 1994;371:606–609. Kimura W, Morikane K, Esaki Y, Chan WC, Pour PM. Histo- logical and biological patterns of microscopic ductal adenocarcinomas detected incidentally at autopsy. Cancer 1998;82:1839–1849. Muscarella P, Knobloch TJ, Ulrich AB et al. Identification and sequencing of the Syrian golden hamster (Mesocricetus auratus) p16(INK4a) and p15(INK4b) cDNAs and their homozygous gene deletion in cheek pouch and pancreatic tumor cells. Gene 2001;278:235–243. Ordonez NG, Balsaver AM, Mackay B. Mucinous islet cell (amphicrine) carcinoma of the pancreas associated with watery diarrhea and hypokalemia syndrome. Hum Pathol 1988;19:1458–1461. Permert J, Ihse I, Jorfeldt L, von Schenck H, Arnquist HJ, Larsson J. Improved glucose metabolism after subtotal pancreatectomy for pancreatic cancer. Br J Surg 1993;80: 1047–1050. Permert J, Ihse I, Jorfeldt L, von Schenck H, Arnqvist HJ, Larsson J. Pancreatic cancer is associated with impaired glucose metabolism. Eur J Surg 1993;159:101–107. Permert J, Larsson J, Westermark GT et al. Islet amyloid polypeptide in patients with pancreatic cancer and diabetes. N Engl J Med 1994;330:313–318. Pour PM, Kazakoff K, Carlson K. Inhibition of streptozotocin-induced islet cell tumors and N- nitrosobis(2-oxopropyl)amine-induced pancreatic ex- ocrine tumors in Syrian hamsters by exogenous insulin. Cancer Res 1990;50:1634–1639. Pour PM, Weide L, Liu G et al. Experimental evidence for the origin of ductal-type adenocarcinoma from the islets of Langerhans. Am J Pathol 1997;150:2167–2180. Pour PM, Schmied BM, Ulrich AB, Friess H, Andren- Sandberg A, Buchler MW. Abnormal differentiation of CHAPTER 44 375 islet cells in pancreatic cancer. Pancreatology 2000;1:110– 116. Pour PM, Standop J, Batra SK. Are islet cells the gatekeepers of the pancreas? Pancreatology 2002;2:440–448. Rosenberg L, Rafaeloff R, Clas D et al. Induction of islet cell differentiation and new islet formation in the hamster: further support for a ductular origin. Pancreas1996;13:38– 46. Schmied B, Liu G, Moyer MP et al. Induction of adenocarci- noma from hamster pancreatic islet cells treated with N- nitrosobis(2-oxopropyl)amine in vitro. Carcinogenesis 1999;20:317–324. Schmied BM, Liu G, Matsuzaki H et al. Differentiation of islet cells in long-term culture. Pancreas 2000;20:337– 347. Standop J, Schneider MB, Ulrich A et al. The pattern of xenobiotic-metabolising enzymes in the human pancreas. J Toxicol Environ Health 2002, in press. Ulrich AB, Schmied BM, Matsuzaki H et al. Increased expres- sion of glutathione S-transferase-pi in the islets of patients with primary chronic pancreatitis but not secondary chron- ic pancreatitis. Pancreas 2001;22:388–394. Yuan S, Rosenberg L, Paraskevas S, Agapitos D, Duguid WP. Transdifferentiation of human islets to pancreatic ductal cells in collagen matrix culture. Differentiation 1996;61: 67–75. PART III 376 The dismal prognosis of pancreatic cancer is mostly due to the fact that this tumor is usually diagnosed at a late stage. There are no specific early symptoms and diag- nostic imaging has limitations. As a result, the disease often eludes detection during its formative stages. Therefore, accurate tools for early diagnosis and screening are particularly important for this tumor. We also need markers that allow estimation of prognosis, disease progression, and treatment response and which help us to select the optimum therapeutic strategy for a patient. Alterations in gene sequences, expression levels, and protein structure or function are used as tumor markers. This field is fast-moving and expanding, but also littered with numerous examples of might-have- beens. Very few markers have passed successfully from the bench to the bedside. In this chapter we highlight the present state of tumor markers in pancreatic cancer and point out developments that may lead to a diagnos- tic breakthrough in the near future. Because 80–90% of tumors of the exocrine pancreas are adenocarcinomas of ductal cell origin, we focus on markers for ductal pancreatic adenocarcinoma. CA-19-9 CA-19-9 is the most frequently used serum-based marker for pancreatic cancer. The protein is a carbohydrate cell-surface antigen (sialylated lacto-N- fucopentose) related to the Lewis blood group sub- stance. It was originally isolated in 1979 as a colorectal cancer-specific antigen and it is found in the normal ep- ithelial cells of the gallbladder, biliary ducts, pancreas, and stomach. The elevation of CA-19-9 in pancreatic and other malignancies is thought to be due to in- creased production and secretion of the antigen from malignant cells. Multiple studies have shown that while elevations in serum CA-19-9 appear useful in the diag- nosis of adenocarcinoma of the upper gastrointestinal tract and in the surveillance of colon cancer, its greatest sensitivity is in the detection of pancreatic adenocarci- noma. To date, CA-19-9 is considered one of the most useful tumor markers for pancreatic malignancies. However, up to 30% of patients with pancreatic cancer do not exhibit elevated serum CA-19-9 levels. The sen- sitivity of the CA-19-9 serum assay ranges between 69 and 93%, the specificity between 46 and 98%. The higher the level of CA-19-9, the greater the sensitivity and specificity of the assay. Elevations in CA-19-9 cor- relate with the degree of tumor differentiation and with the extent of disease. Consequently, CA-19-9 levels are lower in patients with localized disease and this marker is therefore of little use as a screening marker to detect early pancreatic cancers. It has been suggested that very high levels of CA-19-9 indicate unresectable tumors and that the pretreatment CA-19-9 level is a strong pre- dictor of survival. There are conflicting results about whether the response of CA-19-9 to chemotherapy and/or radiotherapy is useful for predicting survival. In addition, CA-19-9 is a useful marker for detecting recurrent disease and can therefore be used for the surveillance of patients after surgery for pancreatic cancer. A general clinical problem is to determine whether a pancreatic mass is due to malignancy or chronic pan- creatitis. Furthermore, if chronic pancreatitis is estab- lished, it is important to know whether there is any sign 377 45 What can be expected from tumor markers in pancreatic cancer? Thomas Seufferlein and Guido Adler of malignant transformation. CA-19-9 is of very limited value in solving this problem, since elevated CA-19-9 levels are also found in benign processes such as acute and chronic pancreatitis, chronic liver disease, and biliary tract disease. Consequently, in patients with suspected pancreatic cancer due to chronic pancreati- tis, the sensitivity and specificity of serum CA-19-9 in the detection of pancreatic cancer were only 44% and 80% respectively. Marked elevations of CA-19-9 are essentially limited to cirrhosis and acute obstructive cholangitis. Biliary obstruction in the absence of cholangitis does not usually produce significant eleva- tions of CA-19-9. The elevated CA-19-9 levels seen with obstructive cholangitis may be due to increased production from the inflamed epithelial cells, along with leakage into the serum due to elevated biliary tract pressure. In the setting of acute inflammatory process- es, serum CA-19-9 values generally return to normal when biliary drainage is achieved and infection re- solves. Thus, an elevated serum CA-19-9 as a marker for malignancy must be interpreted with caution when a pancreatic mass is associated with an inflammatory hepatobiliary process. Genetic markers for the detection of pancreatic cancer in tissues Because of a better understanding of the genetic pro- gression of many common neoplasms, DNA mutations in oncogenes or tumor-suppressor genes are increasing- ly used as genetic markers. Studies in pancreatic cancers and preneoplastic lesions, the so-called pancreatic in- traepithelial neoplasia (PanIn), led to the discovery of specific genetic modifications that occur at early stages of pancreatic carcinogenesis. For example, overexpres- sion of p21 WAF/CIP1 is an early event in precursor lesions, whereas p53 alterations and the loss of DPC4/Smad4 are late events in PanIn development. Ki-ras mutations Activating Ki-ras mutations are the first genetic changes detected in the progression to pancreatic can- cer. They occur in about 30% of lesions that show the earliest stages of histologic disturbance. Therefore, the analysis of Ki-ras mutations has been regarded as a milestone in the early detection of pancreatic cancer. Ki-ras point mutations at codon 12 are also detectable in 75–100% of pancreatic cancer tissues. However, ductal lesions in patients with chronic pancreatitis, and in the normal pancreas also, exhibit Ki-ras mutations without additional indications of neoplastic transfor- mation such as severe dysplasia or mutated P53 pro- tein. Furthermore, Ki-ras mutations are found in benign pancreatic tumors. Thus, Ki-ras as a single marker is not sufficient to establish the diagnosis of pancreatic cancer in a tissue sample. p53 Alterations in p53 are late events in PanIn develop- ment. Overexpression of p53 is almost exclusively found in pancreatic cancers and not in benign pan- creatic tumors. However, only about half of pancreatic cancers exhibit p53 mutations, which limits the value of p53 analysis for the diagnosis of pancreatic cancer. Telomerase Telomerase is a ribonucleoprotein that is involved in telomere maintenance. The enzyme is required for im- mortalization of cells and is expressed by almost every cancer. Telomerase activity has been found in up to 90% of malignant pancreatic tumors but is virtually absent from benign tumors, suggesting that telomerase is activated concomitantly with carcinogenesis. Telom- erase activity could therefore be an interesting marker for pancreatic cancer. However, telomerase assays that determine the precise level of enzyme activity should be used, since low levels of telomerase can be detected in noncancerous tissues leading to false-positive results in less accurate assays. KOC The KOC (KH domain containing protein overex- pressed in cancer) gene is highly overexpressed in pan- creatic cancer. Recent data suggest that KOC is a highly specific and sensitive marker for pancreatic cancer in tissue samples. Mucin family Mucins are heavily glycosylated, high-molecular- weight glycoproteins that play a protective role for ep- ithelial tissues and are possibly involved in the renewal PART III 378 and differentiation of the epithelium, cell adhesion, and cellular signaling. An aberrant expression pattern of mucins can be detected in various malignancies. Mucins may promote the invasive and metastatic po- tential of tumors by contributing to the cell-surface adhesion properties and through morphogenetic signal transduction. MUC-1 has been shown to be overex- pressed in pancreatic adenocarcinomas and PanIns by immunohistochemistry. Other groups have reported that MUC-4 is the only mucin that is differentially expressed at the mRNA level in pancreatic cancers. Expression of MUC-4 is found in up to 89% of pancre- atic cancers and in all PanIn grades, particularly PanIn 3 lesions. However, a few nonneoplastic lesions, in- cluding reactive ducts in chronic pancreatitis, are also MUC-4 positive in immunohistochemistry. Pancreatic cancer markers in serum A highly sensitive and specific marker that is detectable in the serum of patients at risk of developing pancreatic cancer would be ideal for screening. Apart from CA- 19-9, only few such markers have been described. As described above, CA-19-9 is not suitable as an early marker and is not elevated in up to 30% of patients with pancreatic cancer. Apart from proteins, DNA mutations can be detected in serum or plasma samples. The mechanism by which this DNA is released is poorly understood. Ki-ras mutations were found in the plasma of 27% of patients with pancreatic cancer, particularly when distant metastases were present. Such mutations are also detectable in about 5% of patients with chronic pancreatitis. Thus, Ki-ras mutation analysis in serum is specific but has low sensitivity. The epidermal growth factor receptor (EGFR) is overexpressed in the majority of pancreatic cancers. EGFR mRNA is detectable in the peripheral blood of 18% of patients with pancreatic cancer and not in healthy controls. Thus, this marker may be very specific but is not sensitive enough for screening. MUC-4 mRNA can also be detected in peripheral blood mononuclear cells of pancreatic cancer patients, but is undetectable in peripheral blood mononuclear cells of healthy volunteers or patients with chronic pan- creatitis or other cancers. MUC-4 may indeed be useful in differentiating between chronic pancreatitis and pancreatic cancer in patients with a pancreatic mass. Pancreatic juice: the best screening material for pancreatic cancer? Because of the difficulties in obtaining biopsy speci- mens from patients with suspected pancreatic cancer and the low sensitivity of serum-based approaches, much hope has been placed in the analysis of pancreatic juice. Unfortunately, Ki-ras polymerase chain reaction (PCR) of pancreatic juice or bile has a low sensitivity for diagnosing pancreatic cancer. In a prospective trial, codon 12 mutations of the Ki-ras gene were detected in pancreatic juice and bile of 38% of patients with pan- creatic cancer, 8% of patients with chronic pancreati- tis, 18.7% of patients with other malignancies, and 7.3% of patients with benign diseases or normal findings. In different studies, Ki-ras mutations were detected in pancreatic juice of up to 30% of noncancer- ous patients and in more than 60% of patients with benign mucous cell hyperplasia of pancreatic ductal epithelium with chronic inflammation. However, more sensitive and/or quantitative PCR tests may allow differentiation of pancreatic cancer from chronic pan- creatitis. Using quantitative assays such as restriction fragment length polymorphism (RFLP) or hybridiza- tion protection assays, Ki-ras mutations can be detected in up to 84% and 65% of pancreatic cancers respectively. Mutations of p53 in pancreatic juice were detected in 42% of pancreatic cancers. However, no muta- tions were detectable in mucin-producing adenomas or in chronic pancreatitis or normal tissue, making p53 a specific but not very sensitive marker. Combined analysis of Ki-ras and p53 mutations may therefore enhance the genetic diagnosis of pancreatic cancer. Assessing prognosis The most relevant prognostic factors in pancreatic cancer to date are tumor grade, tumor size greater than 45 mm, resection margin involvement, and perineural invasion. Interestingly, in one study loss of DPC4/Smad4 expression in pancreatic cancer corre- lated with resectability and was associated with improved survival after resection, whereas resection did not improve survival in patients whose tumor ex- pressed DPC4/Smad4. Aberrant expression p21 WAF1 , CHAPTER 45 379 [...]... Total No of patients EUS (%) Ultrasound (%) TDM (%) Angiography MRI (%) 5 37 5 30 40 38 25 16 35 25 100 81 80 97 95 87 88 88 66 79 60 — 40 — 55 47 19 — — 68 20 — 80 53 73 75 76 88 — 61 100 — — 80 85 — — — — 81 — — — — — — — 63 — — 256 85 49 69 83 63 MRI, magnetic resonance imaging; TDM, tomodensitometric examination Table 48. 5 Evaluation of invasion of the arterial system by cancer of the pancreas with... (1995) 26 36 32 35 38 39 29 25 22 25 35 85 % 92% 78% 94% — 76% — — 82 % 64% 69% 72% 72% 66% 80 % 74% 82 % 66% 92% 64% 48% 60% T1 T2 T3 N0 N1 24 67 75 79 142 80 81 85 76 81 250 (80 %) 327 (72%) Total Table 48. 3 Evaluation of local extension of cancer of the pancreas with endoscopic ultrasound (EUS) Staging No of patients EUS (%) Ultrasound (%) TDM (%) T staging N staging 82 143 82 68 35 42 44 48 TDM, tomodensitometric... 2003;1 38: 951–955 Slesak B, Harlozinska-Szmyrka A, Knast W et al Tissue polypeptide specific antigen (TPS), a marker for differentiation between pancreatic carcinoma and chronic pancreatitis A comparative study with CA 19–9 Cancer 2000 ;89 : 83 88 381 PART III Trumper L, Menges M, Daus H et al Low sensitivity of the ki-ras polymerase chain reaction for diagnosing pancreatic cancer from pancreatic juice and. .. the reliability of EUS for locoregional staging of pancreatic cancer is 80 – 85 % for tumoral staging and 72–75% for lymph-node staging (Table 48. 1) These results were reported by Rösch et al in 1995 in a series of 250 patients In all these studies, the EUS data have been compared with 403 PART III Table 48. 1 Evaluation of T and N staging by endoscopic ultrasound (EUS) Table 48. 2 Reliability of endoscopic... of both arterial- and venous-phase datasets using a single bolus injection of contrast Arterial-phase images are acquired 25 s after the start of the injection, while portal venous-phase images are obtained at 60 s Therefore high-resolution arterial and venous vascular maps can be created that are comparable to conventional angiography and serve as an essential anatomic map for staging and presurgical... patients with pancreatic carcinoma: diagnostic utility and prognostic significance J Clin Oncol 1999;17:5 78 584 Eskelinen M, Haglund U Developments in serologic detection of human pancreatic adenocarcinoma Scand J Gastroenterol 1999;34 :83 3 84 4 Gress TM, Muller-Pillasch F, Geng M et al A pancreatic can- cer-specific expression profile Oncogene 1996;13: 181 9– 183 0 Halm U, Schumann T, Schiefke I et al Decrease... cell carcinomas of the pancreas: a twenty-year experience Surgery 1 988 ;104:1011–1017 Tsuneo I, Kunihiro M, Hiroshi F et al Radiologic diagnosis of pancreatic carcinoma Semin Surg Oncol 19 98; 15:23–32 CHAPTER 47 Valls C, And a E, Sanchez A Dual-phase helical CT of pancreatic adenocarcinoma: assessment of resectability before surgery Am J Roentgenol 2002;1 78: 821– 82 6 Zeman RK, Cooper C, Zeiberg AS et al... findings, the specificity of EUS for the diagnosis of pancreatic cancer was significantly higher than that of helical CT (88 % vs 41%; P < 0.005) Finally, another study by Mertz et al compared positron emission tomography (PET) for the assessment of locoregional extension of pancreatic carcinoma Sensitivity for diagnosis of pancreatic cancer was 93% for EUS, 87 % for PET, and only 53% for helical CT EUS was more... function and allergies to iodine-based contrast agents that impede the use of CT are probably the main indications for the use of MRI in staging of pancreatic cancer Positron emission tomography Positron emission tomography (PET) with 18Ffluorodeoxyglucose (18F-FDG) is a nonaggressive technique based on the greater incorporation of glucose and its analog 18F-FDG in malignant cells compared 386 with mostly... comprehensive cloning and sequencing method that is used to identify and quantify gene expression, particularly of low-copy-number genes Using a SAGE approach, mesothelin has been identified as a new marker for pancreatic cancer 380 The future of all markers: proteomic pattern analysis? CHAPTER 45 What can be expected from tumor markers in pancreatic cancer? In the foreseeable future CA-1 9-9 will continue . serum CA-1 9-9 levels. The sen- sitivity of the CA-1 9-9 serum assay ranges between 69 and 93%, the specificity between 46 and 98% . The higher the level of CA-1 9-9 , the greater the sensitivity and specificity. signs of al- tered islet cell differentiation, including the formation of intrainsular ductular structures and the expression of tumor-associated antigens CA-1 9-9 , TAG-72, and/ or DU-PAN-2 in islet. antigen (TPS), a marker for differentia- tion between pancreatic carcinoma and chronic pancreati- tis. A comparative study with CA 19–9. Cancer 2000 ;89 : 83 88 . CHAPTER 45 381 Trumper L, Menges M,