help button home button Endocrine Society JCEM
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Reubi, J. C.
Right arrow Articles by Rivier, J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Reubi, J. C.
Right arrow Articles by Rivier, J.
The Journal of Clinical Endocrinology & Metabolism Vol. 88, No. 7 3312-3320
Copyright © 2003 by The Endocrine Society

Expression of CRF1 and CRF2 Receptors in Human Cancers

Jean Claude Reubi, Beatrice Waser, Wylie Vale and Jean Rivier

Division of Cell Biology and Experimental Cancer Research (J.C.R., B.W.), Institute of Pathology, University of Berne, CH-3010 Berne, Switzerland; and The Clayton Foundation Laboratories for Peptide Biology (W.V., J.R.), The Salk Institute, La Jolla, California 92037-1099

Address all correspondence and requests for reprints to: Jean Claude Reubi, M.D., Division of Cell Biology and Experimental Cancer Research, Institute of Pathology, University of Berne, P.O. Box 62, Murtenstrasse 31, CH-3010 Berne, Switzerland. E-mail: reubi{at}pathology.unibe.ch.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Overexpressed peptide receptors in human tumors represent clinically relevant targets for cancer diagnosis and therapy. Corticotropin-releasing factor (CRF) and its receptors have not been known to be involved in human cancer. The aim of the present study was to investigate such possibility by evaluating the expression of CRF1 and CRF2 receptors using in vitro autoradiography with subtype-selective CRF analogs in more than 200 primary human cancer samples. We show that a majority of pituitary adenomas express CRF receptors, often in high amounts. Whereas ACTH-producing adenomas preferentially express CRF1 receptors, nonfunctioning adenomas (gonadotropin-producing and null-cell adenomas) and GH- and TSH-producing adenomas express CRF2 receptors. Furthermore, several central and peripheral nervous system tumors express CRF receptors: medulloblastomas, paragangliomas, neuroblastomas, and some meningiomas express CRF1 receptors, but ependymomas or Ewing sarcomas do not. Insulinomas can also express CRF receptors, whereas ductal pancreatic cancers or prostatic, colorectal, and non-small cell lung cancers lack CRF receptors. In all receptor-positive tumors, the receptors were located on tumor cells. The high incidence of CRF1 or CRF2 receptors in selected human tumors suggests that unlabeled CRF agonists may be evaluated as inhibitors of tumor cell proliferation in cancer therapy, and radiolabeled CRF analogs may be used for cancer diagnosis and/or radiotherapy.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
PEPTIDES ARE CLINICALLY relevant, diagnostically and therapeutically, as ligands to specific receptors. Indeed, selective tumors and their metastases can be precisely localized in patients by means of in vivo peptide receptor scintigraphy (1, 2, 3, 4). Additionally, radiotherapeutic targeting of receptor-expressing tumors with high doses of these radiolabeled peptides is clinically attractive (5, 6, 7).

The clinical applications of small peptides is primarily based on the observation that their receptors are overexpressed in cancers. Such an overexpression of receptors in cancer has been observed in vitro in particular for somatostatin, vasoactive intestinal polypeptide, gastrin-releasing peptide (GRP), cholecystokinin, neurotensin, and neuropeptide Y receptors (8, 9, 10, 11, 12, 13); the respective peptides all belong to a group of brain gut peptides with predominant neurotransmitter function as well as gastrointestinal and endocrine actions. However, in addition to their physiological action, many of these peptides have also been shown to play specific roles in cancers in as much as they have marked effects on tumor cell growth in animal models (14, 15, 16). The high amount of receptors in tumors may be indicative of their pathophysiological relevance in tumor growth regulation (14, 17).

Corticotropin-releasing factor (CRF) is a 41-amino acid-long hypothalamic peptide exerting a wide spectrum of actions at the pituitary and extrapituitary levels (18). It is best known for its role in initiating pituitary-adrenal responses to stress (19). However, CRF also plays a crucial role in the modulation of immune response and has established gastrointestinal, reproductive, and cardiovascular actions (19, 20, 21). Moreover, based on studies in animal tumor models, CRF could be shown to inhibit tumor proliferation (22, 23). All these actions seem to be mediated by specific CRF receptors, CRF1 and CRF2, (24). Although extensive information on CRF receptor distribution is available in rat and mice tissues, in humans, only selected organs have been shown to express CRF receptors (25, 26, 27, 28, 29, 30). Various cancer cell lines such as neuroblastoma, small cell lung cancer, endometrial adenocarcinoma, and melanoma cell lines also express CRF receptors (22, 31, 32, 33, 34). Conversely, CRF receptor expression in primary human tumors is poorly documented, except for ACTH-producing pituitary adenomas and melanomas (33, 35, 36, 37).

In the present study, we, therefore, investigated whether there was a molecular basis for a putative CRF role in human tumors and/or the development of CRF analogs for tumor targeting. In more than 200 human cancers originating from either established physiological CRF target tissues or other tissues, we evaluated the expression at the protein level of the CRF receptor subtypes CRF1 and CRF2 using in vitro receptor autoradiography with subtype-selective analogs.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Aliquots of surgically resected tumors or biopsy specimens submitted for diagnostic histopathological analysis were frozen immediately after surgical resection and stored at -70 C. The following tumors were investigated: ACTH-producing pituitary adenomas (n = 5); prolactin-producing pituitary adenomas (n = 5); GH-producing pituitary adenomas (n = 21); TSH-producing pituitary adenomas (n = 2); gonadotropin-producing pituitary adenomas (n = 8); null-cell adenomas (n = 14); medulloblastomas (n = 8); neuroblastomas (n = 15); ependymomas (n = 6); meningiomas (n = 18); Ewing sarcomas (n = 7); paragangliomas (n = 18); pheochromocytomas (n = 7); exocrine ductal pancreatic carcinomas (n = 10); insulinomas (n = 15); glucagonomas (n = 4); gastrinomas (n = 6); prostate carcinomas (n = 11); non-small cell lung cancer (n = 11); and colon carcinomas (n = 10). The study conformed to the ethical guidelines of the Institute of Pathology and the University of Berne and was approved by its committee.

CRF receptor autoradiography

Twenty-micrometer-thick cryostat sections of the tissue samples were processed for CRF receptor autoradiography as described in detail previously for other peptide receptors (8, 10). The radioligand used was 125I-[Tyr0, Glu1, Nle17]-sauvagine (2000 Ci/mmol; Anawa, Wangen, Switzerland), known as the universal ligand for CRF receptors able to detect CRF1 as well as CRF2 receptors (38). For autoradiography, tissue sections were mounted on precleaned microscope slides and stored at -20 C for at least 3 d to improve adhesion of the tissue to the slide. Sections were then processed according to De Souza and Kuhar (39). They were first preincubated in 0.05 M Trizma buffer (pH 7.4) twice for 15 min at room temperature. The slides were then incubated in a solution containing the same medium as the preincubation solution in which the following compounds were added: 0.1% BSA, 0.1 mM bacitracin, 5 mM MgCl2, 2 mM ethyleneglycotetraacetic acid, 0.3 µM aprotinin, and the radioligand at approximate concentration of 90 pM 125I-[Tyr0, Glu1, Nle17]-sauvagine. The slides were incubated at room temperature with the radioligand for 120 min. To estimate nonspecific binding, paired serial sections were incubated as described above, except that 20 nM unlabeled ligand was added to the medium. To differentiate between CRF1 and CRF2 receptors, the use of subtype-selective CRF analogs, either Stressin1 as CRF1-selective agonist (40) or Astressin2-B as CRF2-selective antagonist (38), were used in competition experiments with the universal CRF radioligand 125I-[Tyr0, Glu1, Nle17]-sauvagine. Moreover, the universal antagonist D-Tyr1-astressin (41) and human urocortin, an endogenous universal CRF agonist for both CRF1 and CRF2 receptors, were tested under the same conditions.

Displacement experiments were performed with 90 pM 125I-[Tyr0, Glu1, Nle17]-sauvagine and increasing amounts of nonradioactive [Tyr0, Glu1, Nle17]-sauvagine, human urocortin, D-Tyr1-astressin, Stressin1, and Astressin2-B to generate competitive inhibition curves on successive sections using the same incubation medium as above. Complete displacement curves performed for all compounds in a series of neoplastic and nonneoplastic tissues led us to conclude that a 20-nM concentration of Stressin1 or Astressin2-B was adequate to evaluate their rank order of potencies in a given tumor and, therefore, to distinguish CRF1 from CRF2 subtype expression in that tumor tissue. On completion of the incubation, the slides were washed five times in ice-cold preincubation solution (pH 7.4) containing 1% BSA and then dried under a stream of cold air, apposed to Biomax MR films (Kodak, Rochester, NY) and exposed for 7 d in x-ray cassettes.

The autoradiograms were quantified using a computer-assisted image processing system, as described previously (8, 10). Tissue standards for iodinated compounds (Amersham, Aylesbury, UK) were used for this purpose. A tissue was defined as receptor positive when the absorbance measured in the total binding section was at least twice that of the nonspecific binding section. The intra- and interobserver reliability of the receptor quantification of the CRF receptor-positive tumors was found to be high (r = 0.98 for both intra- and interobserver reproducibility in the correlation analysis of Pearson). In each experiment, we included as positive controls CRF1-expressing tissues (rat cortex) and CRF2-expressing tissues (rat choroid plexus) (38). Here, we describe the data with predominant expression of receptor subtypes; this is to indicate that if one of the CRF receptor subtypes is heavily expressed in a tumor, it may mask a low amount of the other subtype in this tumor and, therefore, prevent its detection with the present methodology.

In selected cases, immunohistochemical staining for Factor VIII-related antigen (DAKO Corp., Carpinteria, CA) was performed to identify the localization of blood vessels (42).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Table 1Go summarizes the results of the CRF1 and CRF2 receptor incidence and density in various tumors. The pituitary tumors and the tumors of the nervous systems showed the highest incidence of CRF receptors among the different tumor types tested. CRF receptors were also found occasionally in endocrine pancreatic tumors and meningiomas. They were not expressed in exocrine ductal pancreatic carcinomas, ependymomas, Ewing sarcomas, pheochromocytomas, prostate cancers, non-small cell lung cancers, or colon carcinomas (Table 1Go).


View this table:
[in this window]
[in a new window]
  		TABLE 1. Incidence and density of CRF receptor (CRF-R) subtypes in human tumors

 
In terms of receptor subtypes, in pituitary tumors, four of the five ACTH-producing adenomas expressed CRF1 receptors in high density; CRF1 was expressed in low density in one of five prolactinomas, and three other prolactinomas expressed preferentially CRF2, also in low density. All other types of pituitary adenomas expressed preferentially CRF2 receptors: the great majority of GH-producing adenomas, the two TSH-producing adenoma, and the majority of nonfunctioning pituitary adenomas, including both gonadotropin-producing adenomas and null-cell adenomas, expressed CRF2 receptors in moderate to high density (Table 1Go). All CRF receptor-positive tumors of the nervous system expressed the CRF1 subtype. This includes all medulloblastomas, in which CRF1 is found in moderate density, a majority of neuroblastomas with, however, a much lower receptor density; it also includes a majority of paragangliomas with a high density of CRF1 receptors and, occasionally, meningiomas expressing a low CRF1 receptor density. Neuroendocrine pancreatic tumors, in particular insulinomas (6 of 15) and gastrinomas (1 of 6) were shown to express moderate to high amounts of CRF2. One glucagonoma (one of four) expressed a high density of CRF1. From these results, we can see a tendency for CRF2 receptors to be preferentially expressed in endocrine tumors, except for ACTH adenomas, but tumors of the nervous system express preferentially CRF1 receptors. The quantification of the CRF1 and CRF2 receptor autoradiography data shows that a majority of the CRF receptor-expressing tumors express a moderate to high density of receptors. The low receptor level found in meningiomas, prolactinomas, and neuroblastomas appears to be the exception. The receptor density in tumors of a same group can vary up to 10 times from individual to individual (Table 1Go).

To differentiate between CRF1 and CRF2 receptors, the use of subtype-selective CRF analogs, either Stressin1 as CRF1-selective agonist or Astressin2-B as CRF2-selective antagonist, were used in competition experiments in receptor autoradiographical studies with the universal CRF radioligand 125I-[Tyr 0, Glu1, Nle17]-sauvagine. Figure 1Go shows that the rank order of binding affinity of the CRF1- and CRF2-selective analogs obtained in an established CRF1 tissue such as the rat cerebellum (25) is very similar to the rank order of affinity of the same analogs in a human medulloblastoma or paraganglioma. In all cases, there is a high-affinity displacement by the CRF1-selective analog as well as the unlabeled universal CRF ligand, whereas the CRF2-selective analog is inactive. This strongly suggests that the medulloblastoma and paraganglioma are CRF1-expressing tumors. Figure 1Go shows, furthermore, a characteristic rank order of affinity for the CRF1- and CRF2-selective analogs in an established CRF2-expressing tissue, the rat choroid plexus (25). There, a high affinity for the CRF2 analog has been observed, whereas a lower affinity for the CRF1-selective analog is seen. The same pattern is also observed in two human pituitary tumors, a GH-secreting adenoma and a gonadotropin-secreting adenoma, indicating that these tumors also express predominantly CRF2 receptors. Note the very high binding affinity of human urocortin, an endogenous universal CRF ligand, for both CRF1 and CRF2 receptors (Fig. 1Go).



View larger version (31K):
[in this window]
[in a new window]
  		FIG. 1. Competition experiments in CRF1- and CRF2-expressing tissues. CRF1-expressing tissues: rat cerebellum (top left), human medulloblastoma (middle left), and paraganglioma (bottom left). CRF2-expressing tissues: rat choroid plexus (top right), human GH adenoma (middle right), and gonadotropin-secreting adenoma (bottom right). Successive tissue sections were incubated with 90 pM of the radioligand and increasing concentrations of [Tyr0, Glu1, Nle17]-sauvagine (•; univ), Stressin1 ({blacktriangleup}; CRF1-sel), Astressin2-B ({blacklozenge}; CRF2-sel), or human urocortin ({blacksquare}; hUCN). Each graph is a representative example of one tissue. The three graphs on the left show high-affinity displacement of the radioligand by the CRF1-selective Stressin1 but not by the CRF2-selective Astressin2-B. Conversely, the three graphs on the right show high affinity displacement of the radioligand by the CRF2-selective Astressin2-B but not by Stressin1. In all six cases, the unlabeled [Tyr0, Glu1, Nle17]-sauvagine ligand is included as universal ligand. In the two bottom graphs, human urocortin is added as additional universal ligand. Note the very high affinity of urocortin in both cases.

 
Figure 2Go gives representative examples of CRF1- and CRF2-expressing tumors using receptor autoradiography. A CRF1-expressing paraganglioma and a CRF1-expressing ACTH-secreting pituitary adenoma are shown with the characteristic affinity profile for the two subtype-selective analogs. Figure 2Go also illustrates a CRF2-expressing inactive pituitary adenoma and a CRF2-expressing TSH-adenoma with the opposite affinity pattern for the selective analogs. Figure 3Go shows the CRF receptor expression in pancreatic tumors. An insulinoma has CRF2 receptors, but an exocrine pancreatic cancer is CRF receptor negative.



View larger version (95K):
[in this window]
[in a new window]
  		FIG. 2. Receptor autoradiographic demonstration of CRF1 in an ACTH-secreting pituitary adenoma (A–E) and a paraganglioma (F–K) as well as CRF2 in a gonadotropin (L–P)- and TSH-secreting pituitary adenoma (Q–U). Vertical rows: 1 (A, F, L, and Q), Hematoxylin and eosin-stained sections. Bars, 1 mm; 2 (B, G, M, and R), autoradiograms showing total binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine; 3 (C, H, N, and S), autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM of a universal ligand (D-Tyr1-Astressin in C and H, urocortin in N and S); 4 (D, I, O, and T), autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM of the CRF1-selective analog Stressin1; 5 (E, K, P, and U), autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM of the CRF2-selective analog Astressin2-B. High-affinity displacement by Stressin1 is seen in the paraganglioma and ACTH adenoma but not in the two other tumors, and displacement by Astressin2-B is seen in the gonadotropin- and TSH-secreting adenoma but not in the two other tumors.

 


View larger version (82K):
[in this window]
[in a new window]
  		FIG. 3. CRF receptors in pancreatic tumors. A–E, Neuroendocrine pancreatic tumor (insulinoma). F–K, Exocrine ductal pancreatic carcinoma (Ca). A and F, Hematoxylin and eosin-stained sections. Bars, 1 mm. B and G, Autoradiograms showing total binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine. The insulinoma but not the pancreatic carcinoma (Ca) is strongly labeled. C and H, Autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of the universal ligand urocortin (nonspecific binding). D and I, Autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM Stressin1. E and K, Autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM Astressin2-B. The insulinoma has CRF2 receptors, whereas the pancreatic carcinoma has no CRF receptors.

 
In the present study, the localization of the CRF receptors corresponded primarily to tumoral tissue. Figure 4Go shows at high magnification that the CRF1 or CRF2 receptors are expressed by the tumor cells. Moreover, when sections are stained for Factor VIII-related immunoreactivity to visualize blood vessels, it can be observed that the pattern of CRF labeling is distinct from the pattern for tumoral vessel localization; therefore, as seen in a null cell pituitary adenoma in Fig. 4Go, CRF receptors are not predominantly of intratumoral vascular origin but of tumoral origin. Among the 200 tumors analyzed, we were able to identify CRF receptors in tumoral and/or peritumoral vessels in five meningioma samples only. They were all of the CRF2 type. Figure 5Go shows the CRF2 receptor-positive vessels, identified with Factor VIII-related immunoreactivity, in one of the meningiomas.



View larger version (182K):
[in this window]
[in a new window]
  		FIG. 4. Tissue localization of CRF receptors. Left (A–H), High-magnification photographs showing CRF2 (A–D) and CRF1 (E–H) receptors expressed by tumor cells (Tu). A and E, Hematoxylin and eosin-stained sections showing tumor cell nests of a gastrinoma (A) and a paraganglioma (E). Bars, 0.1 mm. B and F, Autoradiograms showing total binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine. The labeling is restricted to the tumor cells (Tu). C and G, Autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM Stressin1. D and H, Autoradiograms showing 125I-[Tyr0, Glu1, Nle17]-sauvagine binding in presence of 20 nM Astressin2-B. Right (I–M), The pattern of CRF receptor localization corresponds to tumor cell localization rather than blood vessel localization in a CRF2-expressing null cell pituitary adenoma. I, Hematoxylin and eosin-stained section showing the pituitary adenoma in the whole section. Bar, 0.1 mm. K, Factor VIII-related immunohistochemistry in an adjacent section showing the tumoral blood vessel localization as black dots or circles. L, Autoradiograms showing total binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine in the whole tumor tissue. M, Autoradiograms showing binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine in presence of 20 nM Astressin2-B. The CRF2 distribution is compatible with the diffuse tumor cell localization but not with the patchy blood vessel pattern.

 


View larger version (53K):
[in this window]
[in a new window]
  		FIG. 5. Vascular CRF2 receptors in tumoral vessels of a meningioma. A, Factor VIII-related immunohistochemistry showing tumoral blood vessels (two of them are identified by arrowheads). B, Autoradiograms showing total binding of 125I-[Tyr0, Glu1, Nle17]-sauvagine. The vessels are strongly labeled (arrowheads). C, Autoradiograms showing binding of 125I-[Tyr0, Glu1, Nle17] in presence of 20 nM Astressin2-B. Complete displacement of the radioligand indicates CRF2 receptors.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
This is the first study reporting the expression of CRF1 and CRF2 receptor proteins in a large number of human cancers of various origins. The study shows not only that several human cancers express a high density of CRF receptors in a high incidence but also that certain tumor types preferentially express CRF1, whereas others express predominantly CRF2 receptors. The extensive expression of CRF receptors in cancer may add a new dimension to the CRF biology, suggesting a potential impact of CRF in cancer research and oncology in addition to its role in stress biology and endocrinology.

In general, the presence of CRF receptors seems to be linked to tumors originating from a CRF-responsive normal tissue. This is true for pituitary adenomas, the great majority of which expresses CRF receptors; it is well established that normal pituitaries can express CRF receptors (39). The above statement is also true for central nervous system tumors: medulloblastomas are tumors originating in the cerebellum, a tissue known to be rich in CRF receptors (39). Also neuroblastomas and paragangliomas, originating from peripheral nervous system tissue, often express CRF receptors. Also, some endocrine pancreatic tumors express CRF receptors, and we have recently shown that urocortin III is present in pancreatic ß-cells and can stimulate insulin and glucagon secretion in vitro and in vivo (Li, C., and W. Vale, personal communication). We do not know very much about the CRF receptor expression of the tissues of origin of other tumors such as meningiomas, pheochromocytomas, Ewing sarcomas, prostate cancers, non-small cell lung cancers, or pancreatic and colon carcinomas. Prostate as well as prostate cancers, though, have been shown to express urocortin (43).

The CRF receptor subtype expressed in these tumors appears to reflect the subtype expressed in the tissue of origin, at least as far as we can judge from the information available. Pituitary corticotrophs express CRF1, and gonadotrophs express CRF2 (Kageyama, K., C. Li, and W. Vale, personal communication); the corresponding ACTH-producing adenomas express CRF1; CRF1 was originally cloned from an ACTH-producing pituitary adenoma (44). Gonadotropin-producing adenomas express CRF2. Null-cell adenomas, which also represent gonadotropin-secreting tumors in the majority of the cases (45), also have CRF2. The cerebellum is an abundant source of CRF1 (25), and all medulloblastomas originating from this tissue have CRF1. Medulloblastomas, which are defined as central primitive neuroectodermal tumors, appear to differ in their molecular and biological characteristics from peripheral primitive neuroectodermal tumors, such as Ewing sarcomas, that do not have CRF receptors. Based on the preceding observations, it is tempting to speculate that human somatotrophs and thyrotrophs may express CRF2 receptors, in analogy to the high expression of CRF2 in GH and TSH adenomas. This, however, remains to be established. To which extent the peripheral nervous system can express CRF1, and, therefore, explain the CRF1 expression in neuroblastomas and paragangliomas, is not known either; it is known only that several of the peripheral actions of CRF are mediated by the peripheral nervous system (20).

The mechanisms controlling the expression of CRF receptors in cancer tissues are not known. These mechanisms are poorly investigated in other peptide receptor systems heavily involved in cancer as well. That neoplastic transformation can lead to a marked overexpression of physiologically occurring peptide receptors in a tissue has been shown previously for somatostatin receptors (17) and has been the molecular basis for clinical applications of somatostatin analogs in oncology, in particular in endocrine gastrointestinal tumors. Conversely, a down-regulation of somatostatin receptors (e.g. sst2) has been observed in exocrine pancreatic cancers (46). A further example is given by the high amount of GRP receptors overexpressed in both breast and prostate cancers (11, 47); breast tumors are originating from breast tissue expressing GRP receptors (47), but prostate carcinomas have their origin in the GRP receptor-negative prostate (11). No general rule as to why a tumor will express a peptide receptor or not can, therefore, be deducted yet from these observations.

Although the tissue resolution of receptor autoradiography is limited, it is evident that the CRF receptor expression is primarily located in the tumor tissue, as shown by the precise matching of the histological tumor distribution and the autoradiographical pattern. It is, however, known that vessels can express CRF receptors of subtype 2 (48, 49). Because vessels are abundantly present in neoplasms, it cannot be excluded that some of the vessels located in CRF receptor-expressing tumors would also express CRF receptors. However, vessels are clearly not the predominant source of CRF receptors in these tumors for the following reasons: 1) The CRF receptor autoradiographic pattern observed in the great majority of the tumors in our study does not correspond to the pattern of vessel localization revealed by Factor VIII-like immunostaining (42) (Fig. 4Go); 2) the vessels would be expected to express predominantly CRF2 (48); and 3) even in CRF receptor-negative tumors with a low background, we were not able to detect much CRF receptors in tumoral or peritumoral vessels. Among all the 200 tumor samples tested, we could identify five meningiomas expressing vascular CRF2 receptors. This indicates that, under particular conditions, tumoral vessels can express measurable levels of CRF2 receptors. This may explain the presence of CRF2 mRNA detected in human vessels in previous reports (48, 49).

The presence of CRF receptors in neoplasms may be propitious for certain clinical applications. A functional CRF receptor in tumors may be used as a target for long-term CRF therapy. Unfortunately, the biology of CRF in human cancer tissue is still poorly understood. There is evidence for an antiproliferative effect of CRF in endometrial cancer cells and W256 rat mammary cancer cells, coupled with a differentiation-inducing effect (22, 23), but there is no information on the CRF capability to inhibit cell proliferation in cell lines derived from other tumor types. It is nevertheless tempting to try to use nonradioactive CRF agonists as a novel long-term strategy to treat the CRF receptor-positive cancer types identified in the present study. Alternatively, the CRF-binding sites in tumors may be used as targets for radiolabeled CRF analogs for the in vivo localization of tumors in the patients and/or the in vivo radiotherapy to selectively destroy tumor tissue. The recent development of CRF1- and CRF2-selective analogs (38, 40) opens the possibility to selectively target the CRF receptor subtype expressed by a given tumor.


   Acknowledgments
 
We thank Dr. A. Schmassmann (Bern and Sursee, Switzerland) for his valuable help with the statistical evaluation.


   Footnotes
 
This work was supported in part by NIH Grant DK26741.

Abbreviations: CRF, Corticotropin-releasing factor; GRP, gastrin-releasing peptide.

Received November 25, 2002.

Accepted April 1, 2003.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Krenning EP, Kwekkeboom DJ, Pauwels S, Kvols LK, Reubi JC 1995 Somatostatin receptor scintigraphy. In: Freeman LM, ed. Nuclear medicine annual 1995. New York: Raven Press; 1–50
  2. Virgolini I, Raderer M, Kurtaran A, Angelberger P, Yang Q, Radosavljevic M, Leimer M, Kaserer K, Li SR, Kornek G, Hübsch P, Niederle B, Pidlich J, Scheithauer W, Valent P 1996 123-I-vasoactive intestinal peptide (VIP) receptor scanning: update of imaging results in patients with adenocarcinomas and endocrine tumors of the gastrointestinal tract. Nucl Med Biol 23:685–692[CrossRef][Medline]
  3. Behr TM, Jenner N, Behe M, Angerstein C, Gratz S, Raue F, Becker W 1999 Radiolabeled peptides for targeting cholecystokinin-B/gastrin receptor-expressing tumors. J Nucl Med 40:1029–1044[Abstract/Free Full Text]
  4. Van de Wiele C, Dumont F, Vanden Broecke R, Oosterlinck W, Cocquyt V, Serreyn R, Peers S, Thornback J, Slegers G, Dierckx RA 2000 Technetium-99m RP527, a GRP analogue for visualisation of GRP receptor-expressing malignancies: a feasibility study. Eur J Nucl Med 27:1694–1699[CrossRef][Medline]
  5. Paganelli G, Zoboli S, Cremonesi M, Bodei L, Ferrari M, Grana C, Bartolomei M, Orsi F, De Cicco C, Macke HR, Chinol M, de Braud F 2001 Receptor-mediated radiotherapy with 90Y-DOTA-D-Phe1-Tyr3-octreotide. Eur J Nucl Med 28:426–434[CrossRef][Medline]
  6. Otte A, Herrmann R, Heppeler A, Behe M, Jermann E, Powell P, Maecke HR, Muller J 1999 Yttrium-90 DOTATOC: first clinical results. Eur J Nucl Med 26:1439–1447[CrossRef][Medline]
  7. Krenning EP, de Jong M, Kooij PP, Breeman WA, Bakker WH, de Herder WW, van Eijck CH, Kwekkeboom DJ, Jamar F, Pauwels S, Valkema R 1999 Radiolabelled somatostatin analogue(s) for peptide receptor scintigraphy and radionuclide therapy. Ann Oncol 10:S23–S29
  8. Reubi JC, Gugger M, Waser B, Schaer JC 2001 Y1-mediated effect of neuropeptide Y in cancer: breast carcinomas as targets. Cancer Res 61:4636–4641[Abstract/Free Full Text]
  9. Reubi JC, Waser B, Schaer JC, Laissue JA 2001 Somatostatin receptor sst1-sst5 expression in normal and neoplastic human tissues using receptor autoradiography with subtype-selective ligands. Eur J Nucl Med 28:836–846[CrossRef][Medline]
  10. Reubi JC, Läderach U, Waser B, Gebbers J-O, Robberecht P, Laissue JA 2000 Vasoactive intestinal peptide/pituitary adenylate cyclase-activating peptide receptor subtypes in human tumors and their tissues of origin. Cancer Res 60:3105–3112[Abstract/Free Full Text]
  11. Markwalder R, Reubi JC 1999 Gastrin-releasing peptide receptors in the human prostate: relation to neoplastic transformation. Cancer Res 59:1152–1159[Abstract/Free Full Text]
  12. Reubi JC, Waser B, Friess H, Büchler MW, Laissue JA 1998 Neurotensin receptors: a new marker for human ductal pancreatic adenocarcinoma. Gut 42:546–550[Abstract/Free Full Text]
  13. Reubi JC, Schaer JC, Waser B 1997 Cholecystokinin(CCK)-A and CCK-B/gastrin receptors in human tumors. Cancer Res 57:1377–1386[Abstract/Free Full Text]
  14. Schally AV 1988 Oncological applications of somatostatin analogs. Cancer Res 48:6977–6985[Medline]
  15. Zia H, Hida T, Jakowlew S, Birrer M, Gozes Y, Reubi JC, Fridkin M, Gozes I, Moody TW 1996 Breast cancer growth is inhibited by vasoactive intestinal peptide (VIP) hybrid, a synthetic VIP receptor antagonist. Cancer Res 56:3486–3489[Abstract/Free Full Text]
  16. Moody TW, Venugopal R, Zia R, Patierno S, Leban JJ, McDermed J 1995 BW2258U89: a GRP antagonist which inhibits small cell lung cancer. Life Sci 56:521–529[CrossRef][Medline]
  17. Reubi JC, Laissue JA 1995 Multiple pathways of somatostatin action in neoplastic disease. Trends Pharmacol Sci 16:110–115[CrossRef][Medline]
  18. Dunn AJ, Berridge CW 1990 Physiological and behavioral responses to corticotropin-releasing factor administration: is CRF a mediator of anxiety or stress responses? Brain Res Brain Res Rev 15:71–100[CrossRef][Medline]
  19. De Souza EB, Grigoriadis DE 1994 Corticotropin-releasing factor: Physiology, pharmacology and role in central nervous system and immune disorders. In: Bloom FE, Kupfer DJ, eds. Psychopharmacology: the fourth generation of progress. New York: Raven Press; 505–517
  20. Parkes DG, Weisinger RS, May CN 2001 Cardiovascular actions of CRH and urocortin: an update. Peptides 22:821–827[CrossRef][Medline]
  21. Bale TL, Giordano FJ, Hickey RP, Huang Y, Nath AK, Peterson KL, Vale WW, Lee KF 2002 Corticotropin-releasing factor receptor 2 is a tonic suppressor of vascularization. Proc Natl Acad Sci USA 99:7734–7739[Abstract/Free Full Text]
  22. Graziani G, Tentori L, Portarena I, Barbarino M, Tringali G, Pozzoli G, Navarra P 2002 CRH inhibits cell growth of human endometrial adenocarcinoma cells via CRH-receptor 1-mediated activation of cAMP-PKA pathway. Endocrinology 143:807–813[Abstract/Free Full Text]
  23. Tjuvajev J, Kolesnikov Y, Joshi R, Sherinski J, Koutcher L, Zhou Y, Matei C, Koutcher J, Kreek MJ, Blasberg R 1998 Anti-neoplastic properties of human corticotropin releasing factor: involvement of the nitric oxide pathway. In Vivo 12:1–10[Medline]
  24. Grigoriadis DE, Lovenberg TW, Chalmers DT, Liaw C, De Souze EB 1996 Characterization of corticotropin-releasing factor receptor subtypes. Ann N Y Acad Sci 780:60–80[Medline]
  25. De Souza EB, Grigoriadis DE 2000 Multiple brain corticotropin-releasing factor receptors and binding protein. In: Quirion R, Björklund A, Hökfelt T, eds. Handbook of chemical neuroanatomy. Vol 16. Amsterdam: Elsevier Science BV; 477–508
  26. Florio P, Franchini A, Reis FM, Pezzani I, Ottaviani E, Petraglia F 2000 Human placenta, chorion, amnion and decidua express different variants of corticotropin-releasing factor receptor messenger RNA. Placenta 21:32–37[CrossRef][Medline]
  27. Kimura Y, Takahashi K, Totsune K, Muramatsu Y, Kaneko C, Darnel AD, Suzuki T, Ebina M, Nukiwa T, Sasano H 2002 Expression of urocortin and corticotropin-releasing factor receptor subtypes in the human heart. J Clin Endocrinol Metab 87:340–346[Abstract/Free Full Text]
  28. Muramatsu Y, Sugino N, Suzuki T, Totsune K, Takahashi K, Tashiro A, Hongo M, Oki Y, Sasano H 2001 Urocortin and corticotropin-releasing factor receptor expression in normal cycling human ovaries. J Clin Endocrinol Metab 86:1362–1369[Abstract/Free Full Text]
  29. Muramatsu Y, Fukushima K, Iino K, Totsune K, Takahashi K, Suzuki T, Hirasawa G, Takeyama J, Ito M, Nose M, Tashiro A, Hongo M, Oki Y, Nagura H, Sasano H 2000 Urocortin and corticotropin-releasing factor receptor expression in the human colonic mucosa. Peptides 21:1799–1809[CrossRef][Medline]
  30. Slominski AT, Botchkarev V, Choudhry M, Fazal N, Fechner K, Furkert J, Krause E, Roloff B, Sayeed M, Wei E, Zbytek B, Zipper J, Wortsman J, Paus R 1999 Cutaneous expression of CRH and CRH-R. Is there a "skin stress response system"? Ann N Y Acad Sci 885:287–311[Abstract/Free Full Text]
  31. Dieterich KD, DeSouza EB 1996 Functional corticotropin-releasing factor receptors in human neuroblastoma cells. Brain Res 733:113–118[CrossRef][Medline]
  32. Dieterich KD, Grigoriadis DE, De Souza EB 1994 Corticotropin-releasing factor receptors in human small cell lung carcinoma cells: radioligand binding, second messenger, and northern blot analysis data. Endocrinology 135:1551–1558[Abstract]
  33. Funasaka Y, Sato H, Chakraborty AK, Ohashi A, Chrousos GP, Ichihashi M 1999 Expression of proopiomelanocortin, corticotropin-releasing hormone (CRH), and CRH receptor in melanoma cells, nevus cells, and normal human melanocytes. J Investig Dermatol Symp Proc 4:105–109[Medline]
  34. Schoeffter P, Feuerbach D, Bobirnac I, Gazi L, Longato R 1999 Functional, endogenously expressed corticotropin-releasing factor receptor type 1 (CRF1) and CRF1 receptor mRNA expression in human neuroblastoma SH-SY5Y cells. Fundam Clin Pharmacol 13:484–489[Medline]
  35. Dieterich KD, Gundelfinger ED, Ludecke DK, Lehnert H 1998 Mutation and expression analysis of corticotropin-releasing factor 1 receptor in adrenocorticotropin-secreting pituitary adenomas. J Clin Endocrinol Metab 83:3327–3331[Abstract/Free Full Text]
  36. Grino M, Guillaume V, Boudouresque F, Margioris AN, Grisoli F, Jaquet P, Oliver C, Conte-Devolx B 1988 Characterization of corticotropin-releasing hormone receptors on human pituitary corticotroph adenomas and their correlation with endogenous glucocorticoids. J Clin Endocrinol Metab 67:279–283[Abstract]
  37. Sakai Y, Horiba N, Sakai K, Tozawa F, Kuwayama A, Demura H, Suda T 1997 Corticotropin-releasing factor up-regulates its own receptor gene expression in corticotropic adenoma cells in vitro. J Clin Endocrinol Metab 82:1229–1234[Abstract/Free Full Text]
  38. Rivier J, Gulyas J, Kirby D, Low W, Perrin MH, Kunitake K, DiGruccio M, Vaughan J, Reubi JC, Waser B, Koerber SC, Martinez V, Wang L, Tache Y, Vale W 2002 Potent and long acid corticotropin releasing factor (CRF) receptor 2 selective peptide competitive antagonists. J Med Chem 45:4737–4747[CrossRef][Medline]
  39. De Souza EB, Kuhar MJ 1986 Corticotropin-releasing factor receptors in the pituitary gland and central nervous system: methods and overview. Methods Enzymol 124:560–590[Medline]
  40. Rivier J, Gulyas J, Kirby D, Kunitake K, DiGruccio M, Donaldson C, Vaughan J, Perrin MH, Koerber SC, Martinez V, Tache Y, Wang L, Rivier C, Vale W 2001 Receptor-selective corticotropin releasing factor analogs. Proc 31st Annual Meeting of the Society for Neuroscience, San Diego, CA, pp 413–417 (Abstract 413.17)
  41. Perrin MH, Sutton SW, Cervini LA, Rivier JE, Vale WW 1999 Comparison of an agonist, urocortin, and an antagonist, astressin, as radioligands for characterization of corticotropin-releasing factor receptors. J Pharmacol Exp Ther 288:729–734[Abstract/Free Full Text]
  42. Denzler B, Reubi JC 1999 Expression of somatostatin receptors in peritumoral veins of human tumors. Cancer 85:188–198[CrossRef][Medline]
  43. Arcuri F, Cintorino M, Florio P, Floccari F, Pergola L, Romagnoli R, Petraglia F, Tosi P, Teresa Del Vecchio M 2002 Expression of urocortin mRNA and peptide in the human prostate and in prostatic adenocarcinoma. Prostate 52:167–172[CrossRef][Medline]
  44. Chen R, Lewis KA, Perrin MH, Vale WW 1993 Expression cloning of a human corticotropin-releasing-factor receptor. Proc Natl Acad Sci USA 90:8967–8971[Abstract/Free Full Text]
  45. Lopes MBS 2000 Tumors of the pituitary gland. In: Fletcher CDM, ed. Diagnostic histopathology of tumors. London: Churchill Livingstone; 931–957
  46. Buscail L, Saint-Laurent N, Chastre E, Vaillant JC, Gespach C, Capella G, Kalthoff H, Lluis F, Vaysse N, Susini C 1996 Loss of sst2 somatostatin receptor gene expression in human pancreatic and colorectal cancer. Cancer Res 56:1823–1827[Abstract/Free Full Text]
  47. Gugger M, Reubi JC 1999 GRP receptors in non-neoplastic and neoplastic human breast. Am J Pathol 155:2067–2076[Abstract/Free Full Text]
  48. Simoncini T, Apa R, Reis FM, Miceli F, Stomati M, Driul L, Lanzone A, Genazzani AR, Petraglia F 1999 Human umbilical vein endothelial cells: a new source and potential target for corticotropin-releasing factor. J Clin Endocrinol Metab 84:2802–2806[Abstract/Free Full Text]
  49. Sanz E, Monge L, Fernandez N, Martinez MA, Martinez-Leon JB, Dieguez G, Garcia-Villalon AL 2002 Relaxation by urocortin of human saphenous veins. Br J Pharmacol 136:90–94[CrossRef][Medline]



This article has been cited by other articles:


Home page
 Am. J. Physiol. Gastrointest. Liver Physiol.Home page
A. A. Teitelbaum, M. G. Gareau, J. Jury, P. C. Yang, and M. H. Perdue
Chronic peripheral administration of corticotropin-releasing factor causes colonic barrier dysfunction similar to psychological stress
Am J Physiol Gastrointest Liver Physiol, September 1, 2008; 295(3): G452 - G459.
[Abstract] [Full Text] [PDF]

Home page
 Hum Reprod UpdateHome page
C. Ferretti, L. Bruni, V. Dangles-Marie, A.P. Pecking, and D. Bellet
Molecular circuits shared by placental and cancer cells, and their implications in the proliferative, invasive and migratory capacities of trophoblasts
Hum. Reprod. Update, March 1, 2007; 13(2): 121 - 141.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
T. Fukuda, K. Takahashi, T. Suzuki, M. Saruta, M. Watanabe, T. Nakata, and H. Sasano
Urocortin 1, Urocortin 3/Stresscopin, and Corticotropin-Releasing Factor Receptors in Human Adrenal and Its Disorders
J. Clin. Endocrinol. Metab., August 1, 2005; 90(8): 4671 - 4678.
[Abstract] [Full Text] [PDF]

Home page
 Nucleic Acids ResHome page
S. Gupta, M. Vingron, and S. A. Haas
T-STAG: resource and web-interface for tissue-specific transcripts and genes
Nucleic Acids Res., July 1, 2005; 33(suppl_2): W654 - W658.
[Abstract] [Full Text] [PDF]

Home page
 JNMHome page
J. C. Reubi, H. R. Macke, and E. P. Krenning
Candidates for Peptide Receptor Radiotherapy Today and in the Future
J. Nucl. Med., January 1, 2005; 46(1_suppl): 67S - 75S.
[Abstract] [Full Text] [PDF]

Home page
 EndocrinologyHome page
B. K. Brar, A. Chen, M. H. Perrin, and W. Vale
Specificity and Regulation of Extracellularly Regulated Kinase1/2 Phosphorylation through Corticotropin-Releasing Factor (CRF) Receptors 1 and 2{beta} by the CRF/Urocortin Family of Peptides
Endocrinology, April 1, 2004; 145(4): 1718 - 1729.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Reubi, J. C.
Right arrow Articles by Rivier, J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Reubi, J. C.
Right arrow Articles by Rivier, J.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals