Department of Oncology, University of Turin, Orbassano, Turin, Italy
3
and of manipulating SSTR signaling as a therapeutic strategy, that will be discussed extensively in the following sections of this book.
methods to identify sstr in tissues
Before discussing the current knowledge on SSTR expression in tumors and other pathologies, a short reappraisal of the different methods available and reported to determine SSTR at the tissue level is mandatory to critically consider the expression data currently available.
Several methods have been used to determine the expression of SSTR in tissue samples. All of them have intrinsically limitations and advantages, and therefore, the most powerful data have been generated by the combination of two or more of them. A comparison of the three most relevant methods is repre-sented in Table 3.1.
SSTR tissue localization had originally been demonstrated by means of binding assays of radiolabeled SS analogues [2–4]. however, this method that has the unique property of tracing the presence of “functionally active” receptors is affected by a scarce reproducibility, is applicable to high‐quality frozen material only, and does not recognize the SSTR subtype expressed, unless using highly subtype‐specific analogues [5].
Since the cloning of the five genes, between 1992 and 1994, several studies detected the specific mRnA expression of SSTR in normal tissue and tumors by means of alternative techniques such as the northern blot [6], in situ hybridization [7, 8], or
tAble 3.1 comparison between different methods of tissue identification of sstr
Method Pros Cons Applicability
MeThodS To IdenTIFY SSTR In TISSUeS 23
reverse transcriptase‐polymerase chain reaction (RT‐PCR), either qualitative or quantitative [9–11]. however, in general terms, mRnA expression from tissue extracts is variably affected by signals determined by SSTR‐expressing cells different from those that represent the target, and the quality of the tissue sample is again a relevant clue for obtaining adequate results.
In parallel with mRnA determination, several groups aimed at the development of SSTR‐specific antibodies. The vast majority of them are polyclonal antisera and have been determined either against the n‐ or the C‐terminus of the protein [12–17]. Most of these antibodies are working nicely on paraffin‐embedded tissue, are raised against all or most of SSTR receptor subtypes (although with variable reliability), are com-mercially available, and, therefore, allowed extensive investigations on large archival case series. Moreover, some monoclonal antibodies have also been developed, with special reference to SSTR subtype 2A [18–21], although not commercially available yet at the time of this manuscript preparation.
Besides research purposes, the advantages of immunohistochemical SSTR detec-tion in the clinical practice include a high cost/benefit ratio, high reproducibility in pathology laboratories worldwide, possibility of tissue localization of tumor cells, recognition of the different SSTR subtypes, and, last but not least, applicability on retrospective archival material. In addition, immunohistochemical methods may be applied to preoperative fine‐needle aspiration of cytological or biopsy material [22], allowing SSTR demonstration in inoperable tumors or offering specific information to diagnostic or therapeutic decisions before surgery (Fig. 3.1). Several studies have also documented that immunohistochemistry correlates with other methods in a
(a) (b)
figure 3.1 SSTR type 2A determination in a cellblock preparation from a liver metastasis of a well‐differentiated neuroendocrine carcinoma of unknown primary. (a) hematoxylin and eosin; (b) immunoperoxidase; (a and b) original magnification 400×. (See insert for color rep-resentation of the figure.)
significant proportion of cases, with only minor discrepancies [10, 13, 18, 21–23].
however, a major caveat of immunohistochemistry for SSTR, with special reference to its use as a diagnostic tool, is represented by the lack of standardization, although some scoring systems have been proposed [24, 25]. In general terms, a fine membranous staining is considered the most specific [26], although a cytoplasmic staining may be a result of receptor internalization, especially in the case of tumors coexpressing both SSTR and their natural ligand or in the cases of patients treated with SS analogues [27].
sstr exPression in humAn mAlignAncies
A range of different tumors overexpress SSTR, as compared to nontransformed cells.
The underlying stimuli that induce this overexpression as well as the functional meaning for the biology of tumor cells have not been conclusively explained. It is possible that the upregulation of SSTR serves as homeostatic growth inhibitory auto-crine/paracrine response to the deregulated tumor cell proliferation [28]. nevertheless, with all the limitations related to the complexity of the role of SSTR in cancer biology, it can be asserted that SSTR and their intracellular signaling pathways should generally be considered as tumor suppressive.
sstr exPression in neuroendocrine neoPlAsms
neuroendocrine neoplasms originate from a normal cell population that is physio-logically a target of SS, thus expressing SSTR. Therefore, the generally high level of SSTR expression in these groups of tumors is not surprising.
A tentative list of all data on SSTR‐positive neuroendocrine neoplasms would greatly fail to be comprehensive. Wide literature data (see for review [29–32]) show that SSTR are highly expressed in pituitary adenomas; neuroendocrine tumors of the pancreas, gastrointestinal tract, and lung; paragangliomas/pheochromocytomas;
Merkel cell carcinomas; neuroblastomas; and medullary thyroid carcinomas. hundreds of such tumors have been analyzed by means of various techniques, including binding assays, immunohistochemistry, and mRnA analysis. A wide heterogeneity in SSTR subtype expression in the different tumor types, in different cases of the same tumor type, and even in different cell populations within individual lesions has been reported.
Such high heterogeneity of SSTR distribution partially explains some discrepancies in the clinical features and response to SS analogue therapy observed in neuroendocrine tumors from various sites. According to such literature data, in most cases, individual tumors coexpress different SSTR subtypes, SSTR type 2 being the most frequently represented in all locations, followed by types 3, 5, and 1. Subtype 4 is generally poorly expressed. higher SSTR expression is generally observed in “well‐differentiated”
tumors although a considerable proportion of positive cases might be observed also in poorly differentiated—highly aggressive—lesions, such as small cell lung cancer [33].
Although poorly elucidated from a biological point of view, hormonal secretion by the
SSTR eXPReSSIon In nonneURoendoCRIne MALIgnAnCIeS 25
tumor is also associated with different patterns of expression of SSTR subtypes, such as in the case of pancreatic insulin‐producing tumors that show a low SSTR type 2 expression [34].
Apart from detailing a prevalence of expression, SSTR determination at the tissue level is potentially a complementary approach to better define the diagnostic and therapeutic strategies in patients affected by neuroendocrine tumors. A recent literature focused on the comparison between SSTR tissue determination (mainly by means of immunohistochem-istry) and in vivo imaging using different SS analogue‐based methodologies [24, 25, 35–37], with in general a good correlation. In a previous paper by our group, a relatively high correlation was observed with the response to SS analogue treatment [24]. Moreover, some studies claimed a prognostic value of SSTR type 2 determination in neuroendocrine tumors of the gastrointestinal tract and pancreas [38–40].
Future studies are therefore needed to validate SSTR testing as a relevant marker of clinical usefulness in neuroendocrine neoplasms.
sstr exPression in nonneuroendocrine mAlignAncies A wide spectrum of solid or hematological malignancies has been demonstrated to variably express SSTR [41, 42]. A variety of carcinomas showed in vivo and tissue localization‐based evidence of SSTR expression: such tumors include, among others, cancers from the breast [43–45], lung [18, 22], kidney [45], pancreatobili-ary tract [46], stomach [47], liver [48], colorectum [49], ovpancreatobili-ary [50], thyroid follic-ular cells [51], and prostate [52–55] (Fig. 3.2). Unpublished data from our group onto cell lines (Table 3.2) are consistent with what was reported on tissues earlier.
In some cases, an antiproliferative activity of SS analogues could be demonstrated
figure 3.2 SSTR type 2A clonal expression in prostatic adenocarcinoma (immunoperoxi-dase; original magnification 400×). (See insert for color representation of the figure.)
in vitro in hormone independent prostate cancer models [56]. SSTR subtypes have also been detected in meningiomas, medulloblastomas, and gliomas, in soft tissue sarcomas, and in malignant melanomas; this distribution correlated with either scintigraphic imaging or in vitro tests on SS analogue response [57–59]. Promising clinical applications of SS analogues have also been reported in lymphohemato-logical malignancies [60] and in thymomas [61].
sstr in nonneoPlAstic diseAses
There is strong evidence that selected nontumoral lesion may also express SSTR. For instance, active granulomas in sarcoidosis express SSTR on epithelioid cells [62], whereas inactive or successfully treated fibrosing granulomas devoid of epithelioid cells lack SSTR. Inflamed joints in active rheumatoid arthritis express SSTR, prefer-entially located in the proliferating synovial vessel [63].
Furthermore, inflammatory bowel disease is characterized by an overexpression of SSTR in the vascular system [64] of the altered parts of the gastrointestinal tract.
Concerning SSTR presence and possible applications of SS analogues in nonneoplas-tic diseases, SS actions in modulating the immunological response and angiogenesis together with the high density of expression of SSTR (viz., type 1 and 2) in the retina represent the baseline of very promising applications for therapy in various retinal diseases, from macular edema or macular degeneration to thyrotoxic orbitopathy, ret-inal ischemic damage, and proliferative diabetic retinopathy [65, 66].
unmet clues
despite the wide body of evidence on the presence and function of SSTR in several neoplastic and nonneoplastic human diseases, several issues still deserve further investigation and elucidation.
tAble 3.2 sstr expression at mrnA level in nonneuroendocrine cell linesa
Cell line derivation SSTR mRnA expressed
MonoMAC Monoblastic leukemia 4
MCF7 Breast cancer 2, 5
T47d Breast cancer, apocrine 2, 5
MdAMB231 Breast cancer 2, 4
CALU‐1 Lung cancer, squamous 3
KATo III gastric cancer 1, 2, 5
hT29 Colon cancer 1, 2, 5
h716 Colon cancer (with neuroendocrine features)
1, 2, 3, 5
Mog UVW glioblastoma 2
a Volante, M., unpublished.
ReFeRenCeS 27
The correct methodology for SSTR determination that could be applied in the clinical practice is not well established, so far. Immunohistochemistry seems the most promising but is still limited by the scarce availability of reliable and clinically validated reagents (mostly related to SSTR subtype 2), as well as by the lack of stan-dardization in the interpretation. Moreover, novel molecules with a wider spectrum of affinity to different SSTR subtypes than those currently available [67] claim a reinterpretation of the data available with a better understanding of coexpression modalities of the different SSTR subtypes, also taking into consideration the capa-bility of different subtype to form functionally active homo‐ or heterodimers that modify significantly the activation of intracellular signaling pathways also due to altered agonist‐induced desensitization [68]. Moreover, the SS/SSTR axis is more complex, due to both other peptides with selective affinity to SSTR such as cor-tistatin [31] and due to the capability of SSTR to heterodimerize with other receptors such as dopamine receptors [69]. In this respect, recent studies claim that the coex-pression of SSTR and dopamine receptors (type 2) might open to novel therapeutic strategies with chimeric molecules [70–72].
references
[1] Krulich, L.; dhariwal, A. P.; McCann, S. M.; et al. endocrinology 1968, 83, 783–790.
[2] Lamberts, S. W.; hofland, L. J.; van Koetsveld, P. M.; et al. Journal of Clinical endocrinology and Metabolism 1990, 71, 566–574.
[3] Papotti, M., Macrí, L.; Bussolati, g.; Reubi, J. C. International Journal of Cancer 1989, 43, 365–369.
[4] Reubi, J. C.; Maurer, R.; von Werder, K.; et al. Cancer Research 1987, 47, 551–558.
[5] Reubi, J. C.; Waser, B.; Schaer, J. C.; Laissue, J. A. european Journal of nuclear Medicine 2001, 28, 836–846.
[6] Kong, h.; dePaoli, A. M.; Breder, C. d.; et al. neuroscience 1994, 59, 175–184.
[7] Reubi, J. C.; Schaer, J. C.; Waser, B.; Mengod, g. Cancer Research 1994, 54, 3455–3459.
[8] Janson, e. T.; gobl, A.; Kalkner, K. M.; oberg, K. Cancer Research 1996, 56, 2561–2565.
[9] Sestini, R.; orlando, C.; Peri, A.; et al. Clinical Cancer Research 1996, 2, 1757–1765.
[10] Papotti, M.; Bongiovanni, M.; Volante, M.; et al. Virchows Archiv 2002, 440, 461–475.
[11] nakayama, Y.; Wada, R.; Yajima, n.; et al. Pancreas 2010, 39, 1147–1154.
[12] helboe, L.; Møller, M.; nørregaard, L.; et al. Brain Research Molecular Brain Research 1997, 49, 82–88.
[13] hofland, L. J.; Liu, Q.; Van Koetsveld, P. M.; et al. Journal of Clinical endocrinology and Metabolism 1999, 84, 775–780.
[14] Janson, e. T. Stridsberg, M.; gobl, A.; et al. Cancer Research 1998, 58, 2375–2378.
[15] Kumar, U.; Sasi, R.; Suresh, S.; et al. diabetes 1999, 48, 77–85.
[16] Reubi, J. C.; Kappeler, A.; Waser, B.; et al. American Journal of Pathology 1998, 153, 233–245.
[17] Schindler, M.; Sellers, L. A.; humphrey, P. P.; emson, P. C. neuroscience 1997, 76, 225–240.
[18] Papotti, M.; Croce, S.; Macri, L.; et al. diagnostic Molecular Pathology 2000, 9, 47–57.
[19] Kuan, C. T.; Wikstrand, C. J.; McLendon, R. e.; et al. hybridoma (Larchmt) 2009, 28, 389–403.
[20] Fischer, T.; doll, C.; Jacobs, S.; et al. Journal of Clinical endocrinology and Metabolism 2008, 93, 4519–4524.
[21] Mundschenk, J.; Unger, n.; Schulz, S.; et al. Journal of Clinical endocrinology and Metabolism 2003, 88, 5150–5157.
[22] Papotti, M.; Croce, S.; Bello, M.; et al. Virchows Archiv 2001, 439, 787–797.
[23] Körner, M.; eltschinger, V.; Waser, B.; et al. American Journal of Surgical Pathology 2005, 29, 1642–1651.
[24] Volante, M.; Brizzi, M. P.; Faggiano, A.; et al. Modern Pathology 2007, 2, 1172–1182.
[25] Miederer, M.; Seidl, S.; Buck, A.; et al. european Journal of nuclear Medicine and Molecular Imaging 2009, 36, 48–52.
[26] Reubi, J. C.; Waser, B.; Liu, Q.; et al. Journal of Clinical endocrinology and Metabolism 2000, 85, 3882–3891.
[27] Reubi, J. C.; Waser, B.; Cescato, R.; et al. Journal of Clinical endocrinology and Metabolism 2010, 95, 2343–2350.
[28] Msaouel, P.; galanis, e.; Koutsilieris, M. expert opinion on Investigational drugs 2009, 18, 1297–1316.
[29] de herder, W. W.; hofland, L. J.; van der Lely, A. J.; Lamberts, S. W. endocrine Related Cancer 2003, 10, 451–458.
[30] Volante, M.; Bozzalla‐Cassione, F.; Papotti M. endocrine Pathology 2004, 15, 275–291.
[31] Volante, M.; Rosas, R.; Allìa, e.; et al. Molecular and Cellular endocrinology 2008, 286, 219–229.
[32] Pelosi, g.; Volante, M.; Papotti, M.; et al. Quarterly Journal of nuclear Medicine and Molecular Imaging 2006, 50, 272–287.
[33] Righi, L.; Volante, M.; Tavaglione, V.; et al. Annals of oncology 2010, 21, 548–555.
[34] Vezzosi, d.; Bennet, A.; Rochaix, P.; et al. european Journal of endocrinology 2005, 152, 757–767.
[35] Müssig, K.; oksüz, M. o., dudziak, K.; et al. hormone and Metabolic Research 2010, 42, 599–606.
[36] Kaemmerer, d.; Peter, L.; Lupp, A.; et al. european Journal of nuclear Medicine and Molecular Imaging 2011, 38, 1659–1668.
[37] Ferone, d.; Pivonello, R.; Kwekkeboom, d. J.; et al. Journal of endocrinological Investigations 2012, 35, 528–534.
[38] Asnacios, A.; Courbon, F.; Rochaix, P.; et al. Journal of Clinical oncology 2008, 26, 963–970.
[39] Kim, h. S.; Lee, h. S.; Kim, W. h. Cancer Research and Treatment 2011, 43, 181–188.
[40] Corleto, V. d.; Falconi, M.; Panzuto, F.; et al. neuroendocrinology 2009, 89, 223–230.
[41] Patel, Y. C. Journal of endocrinological Investigations 1997, 20, 348–367.
ReFeRenCeS 29
[42] hofland, L. J.; Lamberts, S. W. endocrine Reviews 2003, 24, 28–47.
[43] Reubi, C.; gugger, M.; Waser, B. european Journal of nuclear Medicine and Molecular Imaging 2002, 29, 855–862.
[44] Kumar, U.; grigorakis, S. I.; Watt, h. L.; et al. Breast Cancer Research and Treatment 2005, 92, 175–186.
[45] Vikić‐Topić, S.; Raisch, K. P.; Kvols, L. K.; Vuk‐Pavlović, S. Journal of Clinical endocrinology and Metabolism 1995, 80, 2974–2979.
[46] Zhao, B., Zhao, h.; Zhao, n.; et al. Journal of hepato‐Biliary‐Pancreatic Surgery 2002, 9, 497–502.
[47] Miller, g. V.; Farmery, S. M.; Woodhouse, L. F.; Primrose, J. n. British Journal of Cancer 1992, 66, 391–395.
[48] Verhoef, C.; van dekken, h.; hofland, L. J.; et al. digestive Surgery 2008, 25, 21–26.
[49] Qiu, C. Z.; Wang, C.; huang, Z. X.; et al. World Journal of gastroenterology 2006, 12, 2011–2015.
[50] hall, g. h.; Turnbull, L. W.; Richmond, I.; et al. British Journal of Cancer 2002, 87, 86–90.
[51] Klagge, A.; Krause, K.; Schierle, K.; et al. hormone and Metabolic Research 2010, 42, 237–240.
[52] Alonso, o.; gambini, J. P.; Lago, g.; et al. Clinical nuclear Medicine 2011, 36, 1063–1064.
[53] dizeyi, n.; Konrad, L.; Bjartell, A.; et al. Urologic oncology 2002, 7, 91–98.
[54] hansson, J.; Bjartell, A.; gadaleanu, V.; et al. Prostate 2002, 53, 50–59.
[55] halmos, g.; Schally, A. V.; Sun, B.; et al. Journal of Clinical endocrinology and Metabolism 2000, 85, 2564–2571.
[56] Plonowski, A.; Schally, A. V.; nagy, A.; et al. International Journal of oncology 2002, 20, 397–402.
[57] Frühwald, M. C.; Rickert, C. h.; o’dorisio, M. S.; et al. Clinical Cancer Research 2004, 10, 2997–3006.
[58] Florio, T.; Montella, L.; Corsaro, A.; et al. Anticancer Research 2003, 23, 2465–2471.
[59] Arena, S.; Barbieri, F.; Thellung, S.; et al. Journal of neuro‐oncology 2004, 66, 155–166.
[60] Ferone, d.; Resmini, e.; Boschetti, M.; et al. Journal of endocrinological Investigations 2005, 28, S111–117.
[61] Ferone, d.; Montella, L.; de Chiara, A.; et al. Frontiers in Bioscience 2009, 14, 3304–3309.
[62] ten Bokum, A. M.; hofland, L. J.; de Jong, g.; et al. european Journal of Clinical Investigations 1999, 29, 630–636.
[63] Paran, d.; Paran, h. Current opinion in Investigational drugs 2003, 4, 578–582.
[64] Reubi, J. C.; Laissue, J.; Waser, B.; et al. Annals of new York Academy of Sciences 1994, 733, 122–137.
[65] davis, M. I.; Wilson, S. h.; grant, M. B. hormone and Metabolic Research 2001, 33, 295–299.
[66] hernaez‐ortega, M. C.; Soto‐Pedre, e.; Piniés, J. A. diabetes Research and Clinical Practice 2008, 80, e8–10.
[67] Schmid, h. A. Molecular and Cellular endocrinology 2008, 286, 69–74.
[68] Pfeiffer, M.; Koch, T.; Schröder, h.; et al. Journal of Biological Chemistry 2001, 276, 14027–14036.
[69] Ferone, d.; gatto, F.; Arvigo, M.; et al. Journal of Molecular endocrinology 2009, 42, 361–370.
[70] Srirajaskanthan, R.; Watkins, J.; Marelli, L.; et al. neuroendocrinology 2009, 89, 308–314.
[71] diakatou, e.; Kaltsas, g.; Tzivras, M.; et al. endocrine Pathology 2011, 22, 24–30.
[72] Saveanu, A.; Muresan, M.; de Micco, C.; et al. endocrine Related Cancer 2011, 18, 287–300.
Somatostatin Analogues: From Research to Clinical Practice, First Edition. Edited by Alicja Hubalewska-Dydejczyk, Alberto Signore, Marion de Jong, Rudi A. Dierckx, John Buscombe, and Christophe Van de Wiele.
© 2015 John Wiley & Sons, Inc. Published 2015 by John Wiley & Sons, Inc.
AbbreviAtions
NET neuroendocrine tumor
SRS somatostatin receptor scintigraphy SST somatostatin
SSTR somatostatin receptor
Nuclear medicine has always been a powerful tool for the study of biological functions and, more recently, for the molecular and histological characterization of tissues under physiological and pathological conditions. The growing knowledge of the biology of normal and pathological tissues leads to the discovery of several biologically active peptides that mediate their function by binding to external membrane‐bound receptors with high affinity. This property made peptide/receptor complex a potential target to be used for molecular characterization of tissues in vivo.
Peptides have good characteristics as radiopharmaceuticals: they are small in size and have a fast renal clearance and easily penetrate into tissues with consequent low