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Absence of Somatostatin Receptors in Human Exocrine Pancreatic Adenocarcinomas
GASTROENTEROLOGY 1988;95:760-3
Absence of Somatostatin Receptors in Human Exocrine Pancreatic Adenocarcinomas J. C. REDBI, U. HORISBERGER, C. E. ESSED, J. JEEKEL, J. G. H. KLIJN, and S. W. J. LAMBERTS
Sandoz Research Institute Berne, Berne, Switzerland, and Departments of Pathology, Surgery, and Medicine and Rotterdam Radiotherapeutic Institution, Erasmus University, Rotterdam, The Netherlands
Somatostatin receptor frequency was evaluated in 12 human exocrine pancreatic carcinomas taken after surgery. The tumors were analyzed by receptor autoradiography on tissue sections and by in vitro binding techniques on tumor homogenates. None of the tested human pancreatic carcinomas was shown to possess specific somatostatin receptors. In comparison, five single tumors taken from rats transplanted with the rat pancreatic adenocarcinoma CA 20948 were found to contain specific high.affinity somatostatin receptors. Also, human endocrine pancreatic tumors, i.e., two insulinomas, did contain somatostatin receptors under identical experimental conditions. These data confirm previous results with other tumors, documenting the absence of somatostatin receptors in highly malignant human carcinomas. They also may represent an explanation at the molecular level for the lack of therapeutic effect of somatostatin analogues such as SMS 201-995 seen in patients with advanced exocrine pancreatic carcinomas. h e poor prognosis of human pancreatic cancer T and the disappointing results, until now, of various treatment modalities such as surgery, chemotherapy, or radiotherapy (1), make the search for innovative drugs to be used in the treatment of this disease of primary importance. Recently, several reports have been published indicating that certain hormones, such as gastrointestinal hormones, epidermal growth factor, and steroids, are involved in the development and growth of the exocrine pancreas (2). The tetradecapeptide somatostatin and its analogues suppress the secretion of most gastrointestinal hormones (3), some of them being involved in the growth regulation of gastrointestinal tumors. Somatostatin influences not only cell function of the exocrine pancreas but also
deoxyribonucleic acid synthesis and cell growth (4). In experimental animals, receptors for somatostatin have been demonstrated in exocrine pancreatic tissue (5) as well as in endocrine pancreatic cells (6). Therefore, somatostatin might be useful not only in the treatment of endocrine hyperfunctioning pancreatic tumors (7,8), but also in the treatment of the exocrine pancreatic carcinoma, as has been shown by Schally and coworkers in experimental pancreatic tumors in rats and hamsters (9,10). It is known that in various types of somatostatin-responsive tumors, such as growth hormone-producing pituitary adenomas or endocrine gastroenteropancreatic tumors, somatostatin receptors located in the tumors may be the mediators of the therapeutic effect of somatostatin (11-13). We have therefore investigated whether the molecular basis for an efficient treatment with somatostatin analogues exists in human pancreatic adenocarcinomas by using receptor autoradiography to measure the presence of specific somatostatin receptors in these tumors.
Patients and Methods Twelve human pancreatic tumors were obtained at surgery from the University Hospital in Rotterdam. The tumors were all histopathologically diagnosed as primary tumors (n = 9) or metastases (2 from liver, 1 from omentum) of exocrine pancreatic adenocarcinomas. The majority of them were relatively differentiated cases. In addition, two endocrine pancreatic tumors (insulinomas) were also investigated. For comparison, five different tumors from a transplantable rat pancreatic adenocarcinoma (CA 20948) were also added to the study. The same experimental procedure for receptor autoradiography was used with each tumor. The tissues were © 1988 by the American Gastroenterological Association
0016-5085/88/$3.50
September 1988
SOMATOSTATIN RECEPTORS IN PANCREATIC TUMORS
761
Figure 1. Autoradiography of somatostatin receptors in a human pan creatic adenocarcinoma (a-c) and a human insulinoma (d-fl. The radioligand used was 125I_[Leu B • o_Trp22, Tyr 25]-somatost atin 28 . a and d are hematoxylin-stained sections, band e are autoradiograms showing total binding, and c and fare autoradiograms showing nonspecific binding (in presence of 10- 6 M somatostatin 28). Note that only the insulinoma contains som atostatin receptors. Exposition time was 7 days. Bars = 1 mm.
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REUEI ET AL.
immediately put on ice and within $30 min frozen at -80°C. The storage time ranged from 10 days to 5 mo. Tumor samples were between 2 and 20 mm in diameter. Frozen material was cut on a cryostat for autoradiographic visualization of somatostatin receptors as described previously (11,13) using the stable octapeptide 125I-204-090 (a Tyr" analogue of SMS 201-995), or the somatostatin 28 analogue 125I_[Leu8, D_T rp22, Tyr 25]-somatostatin 28.
Results None of the 12 human exocrine pancreatic adenocarcinomas displayed somatostatin receptors, neither with 1251_[Leu8, D_Trp22, Tyr25]-somatostatin 28 nor with 1251-204-090 ligands, but under the same experimental conditions two human insulinomas were somatostatin receptor-positive. Furthermore, all five rat pancreatic tumors were shown to bear specific somatostatin receptors, both with 1251_[Leu8, D-Trp22, Tyr 25]-somatostatin 28 and 1251_204_090 ligands. Biochemical and pharmacologic characterization of the rat tumors revealed saturable, highaffinity binding sites (KD = 2.S nM) with pharmacologic specificity for somatostatin. Figures la-lc show one example of a somatostatin receptornegative human pancreatic adenocarcinoma, whereas Figures ld-lf show a somatostatin receptorpositive human insulinoma. In all receptor-positive tumors, somatostatin receptors are restricted to tumor cell areas, as already described for various human tumors containing such receptors (11-13). As a positive control for the biologic quality of the tissues examined, epidermal growth factor receptors, known to occur in pancreatic carcinomas (14), were detected in 6 of 8 cases of somatostatin receptornegative human pancreatic adenocarcinomas using similar autoradiographic techniques.
Discussion The present results demonstrate the absence of somatostatin receptors in all tested human exocrine pancreatic adenocarcinomas. These data contrast with the description of somatostatin receptors in five tumors from rats transplanted with the pancreatic carcinoma CA 20948 as well as in the normal rat exocrine pancreas (S). Unfortunately, no detailed somatostatin receptor analysis of the normal human pancreas is available at the present time. The absence of somatostatin receptors in human exocrine pancreatic tumors may be related to the high malignancy state of most of these human tumors. Indeed, there is evidence that somatostatin receptors are rather localized on slow-growing, highly differentiated tumor types. First, they are present in most growth hormone-producing pituitary adenomas (12). Second, they are present in a
GASTROENTEROLOGY Vol. 95, No.3
large proportion of differentiated glia-derived brain tumors, such as astrocytomas and oligodendrogliomas, whereas they are absent in the poorly differentiated glioblastomas (is). Furthermore, differentiated human endocrine tumors of the pancreas do possess, in many instances, somatostatin receptors (13), as confirmed by the present study. As there is increasing evidence that the beneficial effect in vivo of somatostatin analogues in various tumors, such as growth hormone-producing pituitary adenomas or various hormone-producing gastrointestinal tumors, may be related to the presence of specific somatostatin receptors located at the tumor site (11-13), it is relevant to question whether somatostatin analogues may be expected to be effective in somatostatin receptor-negative tumors such as human exocrine pancreatic adenocarcinomas. Our results imply that no direct interaction of somatostatin with the tumor itself will take place. It should, however, be pointed out that somatostatin has a diffuse, broad range of actions throughout the body, inhibiting the secretion of growth hormone, production of somatomedin, and secretion of thyroid-stimulating hormone, gastroenteropancreatic hormones, and possibly of other growth factors. An indirect mechanism of action of somatostatin, as postulated in certain animal tumors (7), cannot therefore be excluded. However, in a preliminary study involving 11 patients with extensively metastasized pancreatic carcinomas, we did not observe an objective remission or stabilization during chronic therapy for 3-9 mo with 200 J-Lg SMS 20199S t.i.d, The present data therefore indicate that somatostatin analogues are unlikely to hold great therapeutic promise in the treatment of patients with pancreatic adenocarcinomas, probably because such tumors lack somatostatin receptors mediating the drug action locally.
References 1. O'Connell MJ. Current status of chemotherapy for advanced pancreatic and gastric cancer. J CUn Oncol 1985;3:1032-9. 2. Johnson LR. Effects of gastrointestinal hormones on pancreatic growth. Cancer 1983;47:1640-5. 3. Adrian TE, Barnes AJ, Long RG, et al. The effect of somatostatin analogues on secretion of growth, pancreatic and gastrointestinal hormones in man. J CUn Endocrinol Metab 1981;53:675-81. 4. Morisset J. Somatostatin: a potential antigrowth factor for the exocrine pancreas. Regul Pept 1984;10:11-22. 5. Sakamoto C, Goldfine ID, Williams JA. The somatostatin receptor on isolated pancreatic acinar cell plasma membranes. Identification of subunit structure and direct regulation by cholecystokinin. J Biol Chern 1984;259:9623-7. 6. Reubi Je, Rivier J, Perrin M, Brown M, Vale W. Specific high affinity binding sites for somatostatin-28 on pancreatic f3-
September 1988
cells: differences with brain somatostatin receptors. Endocrinology 1982;110:1049-51. 7. Reubi [C. A somatostatin analogue inhibits chondrosarcoma and insulinoma tumor growth. Acta Endocrinol 1985;109: 108-14.
8. Ch'ng JLC, Anderson JV, Williams SJ, Carr DH, Bloom SR. Remission of symptoms during long term treatment of metastatic pancreatic endocrine tumours with long acting soinatostatin analogue. Br Med J 1986;292:981-2. 9. Redding TM, Schally AV. Inhibition of growth of pancreatic carcinomas in animal models by analogues of hypothalamic hormones. Proc Nat! Acad Sci USA 1984;81:248-52. 10. Paz-Bouza JI, Redding TW, Schally AV. Treatment of nitrosamine-induced pancreatic tumors in hamsters with analogs of somatostatin and luteinizing hormone-releasing hormone. Proc Natl Acad Sci USA 1987;84:1112-6. 11. Reubi JC, Maurer R, von Werder K, Torhorst J, Klijn JGM, Lamberts SWJ. Somatostatin receptors in human endocrine tumors. Cancer Res 1987;47:551-8. 12. Reubi JC, Heitz PU, Landolt AM. Visualization of somato-
SOMATOSTATIN RECEPTORS IN PANCREATIC TUMORS
763
statin receptors and correlation with immunoreactive GH and PRL in human pituitary adenomas. Evidence for different tumor subclasses. J Clin Endocrinol Metab 1987;65:65-73. 13. Reubi JC, Hackl WH, Lamberts SWJ. Hormone-producing gastrointestinal tumors contain high density of somatostatin receptors. J Clin Endocrinol Metab 1987;65:1127-34. 14. Korc M, Meltzer PS, Trent J; Enhanced expression of the receptor for epidermal growth factor correlates with alterations of the short arm of chromosome 7 in human pancreatic cancer. Dig Dis Sci 1985;30:978. 15. Reubi JC, Lang W, Maurer R, Koper JW, Lamberts SWJ. Distribution and biochemical characterization of somatostatin receptors in tumors of the human central nervous system. Cancer Res 1987;47:5758-64.
Received January 12, 1988. Accepted April 1, 1988. Address requests for reprints to: J.C. Reubi, M.D., Sandoz Research Institute Berne, P.O. Box 2173, CH 3001 Berne, Switzerland.
Absence of Somatostatin Receptors in Human Exocrine Pancreatic Adenocarcinomas J. C. REDBI, U. HORISBERGER, C. E. ESSED, J. JEEKEL, J. G. H. KLIJN, and S. W. J. LAMBERTS
Sandoz Research Institute Berne, Berne, Switzerland, and Departments of Pathology, Surgery, and Medicine and Rotterdam Radiotherapeutic Institution, Erasmus University, Rotterdam, The Netherlands
Somatostatin receptor frequency was evaluated in 12 human exocrine pancreatic carcinomas taken after surgery. The tumors were analyzed by receptor autoradiography on tissue sections and by in vitro binding techniques on tumor homogenates. None of the tested human pancreatic carcinomas was shown to possess specific somatostatin receptors. In comparison, five single tumors taken from rats transplanted with the rat pancreatic adenocarcinoma CA 20948 were found to contain specific high.affinity somatostatin receptors. Also, human endocrine pancreatic tumors, i.e., two insulinomas, did contain somatostatin receptors under identical experimental conditions. These data confirm previous results with other tumors, documenting the absence of somatostatin receptors in highly malignant human carcinomas. They also may represent an explanation at the molecular level for the lack of therapeutic effect of somatostatin analogues such as SMS 201-995 seen in patients with advanced exocrine pancreatic carcinomas. h e poor prognosis of human pancreatic cancer T and the disappointing results, until now, of various treatment modalities such as surgery, chemotherapy, or radiotherapy (1), make the search for innovative drugs to be used in the treatment of this disease of primary importance. Recently, several reports have been published indicating that certain hormones, such as gastrointestinal hormones, epidermal growth factor, and steroids, are involved in the development and growth of the exocrine pancreas (2). The tetradecapeptide somatostatin and its analogues suppress the secretion of most gastrointestinal hormones (3), some of them being involved in the growth regulation of gastrointestinal tumors. Somatostatin influences not only cell function of the exocrine pancreas but also
deoxyribonucleic acid synthesis and cell growth (4). In experimental animals, receptors for somatostatin have been demonstrated in exocrine pancreatic tissue (5) as well as in endocrine pancreatic cells (6). Therefore, somatostatin might be useful not only in the treatment of endocrine hyperfunctioning pancreatic tumors (7,8), but also in the treatment of the exocrine pancreatic carcinoma, as has been shown by Schally and coworkers in experimental pancreatic tumors in rats and hamsters (9,10). It is known that in various types of somatostatin-responsive tumors, such as growth hormone-producing pituitary adenomas or endocrine gastroenteropancreatic tumors, somatostatin receptors located in the tumors may be the mediators of the therapeutic effect of somatostatin (11-13). We have therefore investigated whether the molecular basis for an efficient treatment with somatostatin analogues exists in human pancreatic adenocarcinomas by using receptor autoradiography to measure the presence of specific somatostatin receptors in these tumors.
Patients and Methods Twelve human pancreatic tumors were obtained at surgery from the University Hospital in Rotterdam. The tumors were all histopathologically diagnosed as primary tumors (n = 9) or metastases (2 from liver, 1 from omentum) of exocrine pancreatic adenocarcinomas. The majority of them were relatively differentiated cases. In addition, two endocrine pancreatic tumors (insulinomas) were also investigated. For comparison, five different tumors from a transplantable rat pancreatic adenocarcinoma (CA 20948) were also added to the study. The same experimental procedure for receptor autoradiography was used with each tumor. The tissues were © 1988 by the American Gastroenterological Association
0016-5085/88/$3.50
September 1988
SOMATOSTATIN RECEPTORS IN PANCREATIC TUMORS
761
Figure 1. Autoradiography of somatostatin receptors in a human pan creatic adenocarcinoma (a-c) and a human insulinoma (d-fl. The radioligand used was 125I_[Leu B • o_Trp22, Tyr 25]-somatost atin 28 . a and d are hematoxylin-stained sections, band e are autoradiograms showing total binding, and c and fare autoradiograms showing nonspecific binding (in presence of 10- 6 M somatostatin 28). Note that only the insulinoma contains som atostatin receptors. Exposition time was 7 days. Bars = 1 mm.
762
REUEI ET AL.
immediately put on ice and within $30 min frozen at -80°C. The storage time ranged from 10 days to 5 mo. Tumor samples were between 2 and 20 mm in diameter. Frozen material was cut on a cryostat for autoradiographic visualization of somatostatin receptors as described previously (11,13) using the stable octapeptide 125I-204-090 (a Tyr" analogue of SMS 201-995), or the somatostatin 28 analogue 125I_[Leu8, D_T rp22, Tyr 25]-somatostatin 28.
Results None of the 12 human exocrine pancreatic adenocarcinomas displayed somatostatin receptors, neither with 1251_[Leu8, D_Trp22, Tyr25]-somatostatin 28 nor with 1251-204-090 ligands, but under the same experimental conditions two human insulinomas were somatostatin receptor-positive. Furthermore, all five rat pancreatic tumors were shown to bear specific somatostatin receptors, both with 1251_[Leu8, D-Trp22, Tyr 25]-somatostatin 28 and 1251_204_090 ligands. Biochemical and pharmacologic characterization of the rat tumors revealed saturable, highaffinity binding sites (KD = 2.S nM) with pharmacologic specificity for somatostatin. Figures la-lc show one example of a somatostatin receptornegative human pancreatic adenocarcinoma, whereas Figures ld-lf show a somatostatin receptorpositive human insulinoma. In all receptor-positive tumors, somatostatin receptors are restricted to tumor cell areas, as already described for various human tumors containing such receptors (11-13). As a positive control for the biologic quality of the tissues examined, epidermal growth factor receptors, known to occur in pancreatic carcinomas (14), were detected in 6 of 8 cases of somatostatin receptornegative human pancreatic adenocarcinomas using similar autoradiographic techniques.
Discussion The present results demonstrate the absence of somatostatin receptors in all tested human exocrine pancreatic adenocarcinomas. These data contrast with the description of somatostatin receptors in five tumors from rats transplanted with the pancreatic carcinoma CA 20948 as well as in the normal rat exocrine pancreas (S). Unfortunately, no detailed somatostatin receptor analysis of the normal human pancreas is available at the present time. The absence of somatostatin receptors in human exocrine pancreatic tumors may be related to the high malignancy state of most of these human tumors. Indeed, there is evidence that somatostatin receptors are rather localized on slow-growing, highly differentiated tumor types. First, they are present in most growth hormone-producing pituitary adenomas (12). Second, they are present in a
GASTROENTEROLOGY Vol. 95, No.3
large proportion of differentiated glia-derived brain tumors, such as astrocytomas and oligodendrogliomas, whereas they are absent in the poorly differentiated glioblastomas (is). Furthermore, differentiated human endocrine tumors of the pancreas do possess, in many instances, somatostatin receptors (13), as confirmed by the present study. As there is increasing evidence that the beneficial effect in vivo of somatostatin analogues in various tumors, such as growth hormone-producing pituitary adenomas or various hormone-producing gastrointestinal tumors, may be related to the presence of specific somatostatin receptors located at the tumor site (11-13), it is relevant to question whether somatostatin analogues may be expected to be effective in somatostatin receptor-negative tumors such as human exocrine pancreatic adenocarcinomas. Our results imply that no direct interaction of somatostatin with the tumor itself will take place. It should, however, be pointed out that somatostatin has a diffuse, broad range of actions throughout the body, inhibiting the secretion of growth hormone, production of somatomedin, and secretion of thyroid-stimulating hormone, gastroenteropancreatic hormones, and possibly of other growth factors. An indirect mechanism of action of somatostatin, as postulated in certain animal tumors (7), cannot therefore be excluded. However, in a preliminary study involving 11 patients with extensively metastasized pancreatic carcinomas, we did not observe an objective remission or stabilization during chronic therapy for 3-9 mo with 200 J-Lg SMS 20199S t.i.d, The present data therefore indicate that somatostatin analogues are unlikely to hold great therapeutic promise in the treatment of patients with pancreatic adenocarcinomas, probably because such tumors lack somatostatin receptors mediating the drug action locally.
References 1. O'Connell MJ. Current status of chemotherapy for advanced pancreatic and gastric cancer. J CUn Oncol 1985;3:1032-9. 2. Johnson LR. Effects of gastrointestinal hormones on pancreatic growth. Cancer 1983;47:1640-5. 3. Adrian TE, Barnes AJ, Long RG, et al. The effect of somatostatin analogues on secretion of growth, pancreatic and gastrointestinal hormones in man. J CUn Endocrinol Metab 1981;53:675-81. 4. Morisset J. Somatostatin: a potential antigrowth factor for the exocrine pancreas. Regul Pept 1984;10:11-22. 5. Sakamoto C, Goldfine ID, Williams JA. The somatostatin receptor on isolated pancreatic acinar cell plasma membranes. Identification of subunit structure and direct regulation by cholecystokinin. J Biol Chern 1984;259:9623-7. 6. Reubi Je, Rivier J, Perrin M, Brown M, Vale W. Specific high affinity binding sites for somatostatin-28 on pancreatic f3-
September 1988
cells: differences with brain somatostatin receptors. Endocrinology 1982;110:1049-51. 7. Reubi [C. A somatostatin analogue inhibits chondrosarcoma and insulinoma tumor growth. Acta Endocrinol 1985;109: 108-14.
8. Ch'ng JLC, Anderson JV, Williams SJ, Carr DH, Bloom SR. Remission of symptoms during long term treatment of metastatic pancreatic endocrine tumours with long acting soinatostatin analogue. Br Med J 1986;292:981-2. 9. Redding TM, Schally AV. Inhibition of growth of pancreatic carcinomas in animal models by analogues of hypothalamic hormones. Proc Nat! Acad Sci USA 1984;81:248-52. 10. Paz-Bouza JI, Redding TW, Schally AV. Treatment of nitrosamine-induced pancreatic tumors in hamsters with analogs of somatostatin and luteinizing hormone-releasing hormone. Proc Natl Acad Sci USA 1987;84:1112-6. 11. Reubi JC, Maurer R, von Werder K, Torhorst J, Klijn JGM, Lamberts SWJ. Somatostatin receptors in human endocrine tumors. Cancer Res 1987;47:551-8. 12. Reubi JC, Heitz PU, Landolt AM. Visualization of somato-
SOMATOSTATIN RECEPTORS IN PANCREATIC TUMORS
763
statin receptors and correlation with immunoreactive GH and PRL in human pituitary adenomas. Evidence for different tumor subclasses. J Clin Endocrinol Metab 1987;65:65-73. 13. Reubi JC, Hackl WH, Lamberts SWJ. Hormone-producing gastrointestinal tumors contain high density of somatostatin receptors. J Clin Endocrinol Metab 1987;65:1127-34. 14. Korc M, Meltzer PS, Trent J; Enhanced expression of the receptor for epidermal growth factor correlates with alterations of the short arm of chromosome 7 in human pancreatic cancer. Dig Dis Sci 1985;30:978. 15. Reubi JC, Lang W, Maurer R, Koper JW, Lamberts SWJ. Distribution and biochemical characterization of somatostatin receptors in tumors of the human central nervous system. Cancer Res 1987;47:5758-64.
Received January 12, 1988. Accepted April 1, 1988. Address requests for reprints to: J.C. Reubi, M.D., Sandoz Research Institute Berne, P.O. Box 2173, CH 3001 Berne, Switzerland.