Antitumor effect of in vivo somatostatin receptor subtype 2 gene transfer in primary and metastatic pancreatic cancer models

Our previous studies conducted in pancreatic cancer models established in nude mice and hamsters revealed that cloned somatostatin receptor subtype 2 (sst2) gene expression induced both antioncogenic and local antitumor bystander effects in vivo. In the present study, in vivo gene transfer of sst2 was investigated in two transplantable models of primary and metastatic pancreatic carcinoma developed in hamsters. LacZ reporter or mouse sst2 genes were expressed by means of two different delivery agents: an adenoviral vector and a synthetic polycationic carrier [linear polyethylenimine (PEI)]. sst2 was injected into either exponentially growing pancreatic primary tumors or hepatic metastases, and then transgene expression and tumor progression were investigated 5-6 days after gene transfer. Molecular mechanisms involved in the inhibition of tumor growth were also analyzed. Both adenovirus- and PEI-mediated in vivo gene transfer in primary pancreatic tumors induced an increase of beta-galactosidase activity and expression of sst2 transgene nRNA (100% and 86% of tumors for adenovirus and PEI vector, respectively). Adenoviral vector-based sst2 gene transfer resulted in significant reduction of pancreatic tumor growth (P < 0.05). Using PEI vector, both pancreatic primary tumor growth and metastatic tumor growth were also significantly slackened as compared with both LacZ-treated and untreated control groups (P < 0.02). Moreover, the proliferative index decreased significantly (P < 0.005), whereas apoptosis increased (P < 0.005) in tumors transferred with sst2 gene. The increase of apoptosis correlated with an activation of the caspase-3 and poly(ADP-ribose) polymerase pathways. We concluded that in both primary and metastatic pancreatic cancer models, the synthetic gene delivery system can achieve in vivo sst2 gene transfer and results in a significant antitumor effect characterized by an increase of apoptosis and an inhibition of cell proliferation. This new strategy of gene therapy allows the restoration of expression of an antioncogenic molecule and could be promising for the treatment of advanced pancreatic cancer.     

Somatostatin receptors in non-neuroendocrine malignancies: the potential role of somatostatin analogs in solid tumors

Somatostatin receptors (sstrs) are G-protein-coupled receptors that mediate various physiological effects when activated by the neuropeptide somatostatin or its synthetic analogs. In addition to the well-documented antisecretory effects of sstr(2)-preferential somatostatin analogs octreotide and lanreotide, ligand binding to sstr initiates an inhibitory action on tumor growth. This effect may result from both indirect actions (suppression of growth factors and growth-promoting hormones [e.g., GH/IGF-1 axis] and inhibition of angiogenesis) and direct actions (activation of antigrowth activities [e.g., apoptosis]). As solid tumor cells express multiple sstrs, there is a rationale to evaluate the potential antitumor effects of pasireotide (SOM230), a multireceptor-targeted somatostatin analog with high binding affinity for sstr(1-3) and sstr(5). Pasireotide reduces systemic IGF-1 levels more potently than currently available somatostatin analogs and has been well tolerated in clinical trials.     

生长抑素-细胞毒素靶向作用的研究进展

生长抑素受体广泛表达于各种肿瘤组织。生长抑素及其类似物与毒素或化疗药物偶联,可以选择性结合到肿瘤细胞表面的特异性生长抑素受体上,毒素内化进入靶细胞。其主要优点在于能减轻常规放、化疗的毒副反应并增强抗癌效应。     

Immunohistochemical determination of five somatostatin receptors in meningioma reveals frequent overexpression of somatostatin receptor subtype sst2A.

Meningioma is one of a variety of human tumors that exhibit a very high density of somatostatin receptors and in many cases show a true positive somatostatin receptor scintigraphy. However, the level of expression of individual somatostatin receptor proteins in meningioma has not been investigated. We have recently developed a panel of somatostatin receptor subtype-specific antibodies that effectively stain formalin-fixed, paraffin-embedded tumor tissue (S. Schulz et al., Clin. Cancer Res., 4: 2047-2052, 1998). In the present study, we have used these antibodies to determine the somatostatin receptor status of 40 randomly selected meningiomas. Immunoreactive staining for all somatostatin receptors was clearly located at the plasma membrane of the tumor cells and completely blocked with antigenic peptide. The vast majority of tumors (29 cases; 70%) were positive for sst2A immunoreactivity; among these, 20 (69%) tumors showed high levels of sst2A immunoreactivity. In contrast, all other somatostatin receptors were only detected sporadically, and none of these cases revealed a particularly strong staining. However, it is uncertain to what extent somatostatin receptor-immunoreactive staining intensity may translate into somatostatin receptor protein expression on the tumor cells. Therefore, in a prospective study, 16 surgically removed meningiomas were collected, and the level of sst2A expression was determined using Western blot analysis. Whereas sst2A was readily detectable as a broad band migrating at Mr 70,000 in 12 (75%) of these tumors, 8 tumors (50%) showed particularly high levels of immunoreactive sst2A receptors. There was an excellent correlation (P < 0.001) between the level of sst2A protein expression detected in Western blots and the sst2A- immunoreactive staining seen in tissue sections. Thus, the frequent overexpression of the sst2A receptor may explain the high tracer uptake often observed in meningioma patients during somatostatin receptor scintigraphy. Moreover, this simple immunohistochemical method could prove useful in identifying those cases of recurrent disease that may possibly respond to therapy with sst2-selective agonists.     

Long-term octreotide treatment of metastatic carcinoid tumor

The optimal dosage of somatostatin analogs for the long-term control ofcarcinoid tumors has not yet been established. Receptor alterations inducedduring long term treatment with somatostatin analogs have lead to consecutivedrug dosage increases in order to control carcinoid disease. In this report,we describe the rapid and effective control of tumor in a patient withmetastatic carcinoid treated for nine years with a single daily dose ofoctreotide based on tumor marker levels. Tumoral somatostatin receptor (sst)subtype analysis by RT-PCR amplification showed the expression ofsst₂ subtype only. We suggest that a single daily dose ofoctreotide strictly related to tumor marker secretion, could have played arole in the effective long-term therapy by avoiding the phenomenon ofsomatostatin receptor desensitisation. Furthermore, the exclusive presence ofsst₂ subtype supports the high affinity of octreotide to tumoralcells.     

Inhibition of growth of MX-1, MCF-7-MIII and MDA-MB-231 human breast cancer xenografts after administration of a targeted cytotoxic analog of somatostatin, AN-238.

Since somatostatin (sst) receptors are expressed in a high percentage of human breast cancers, we studied the effects of a targeted cytotoxic somatostatin analog (AN-238) formed by linking the highly active doxorubicin (DOX) derivative 2-pyrrolino-DOX (AN-201) to octapeptide RC-121 (D-Phe-Cys-Tyr-D-Trp-Lys-Val-Cys-Thr-NH(2)) in 3 human breast cancer models. The models included estrogen-independent MDA-MB-231 and MX-1 and estrogen-sensitive MCF-7-MIII tumors. Nude mice bearing xenografts of these cancers were injected i.v. with 250 nmol/kg doses of cytotoxic radical AN-201, cytotoxic analog AN-238 or the unconjugated mixture of AN-201 and sst analog RC-121. Significant inhibition of growth of MDA-MB-231, MX-1 and MCF-7-MIII tumors was observed 1 week after injection of a single dose of cytotoxic analog AN-238. The volume of MDA-MB-231 tumors remained significantly decreased 3 weeks after treatment. The volumes and weights of MCF-7-MIII tumors continued to be significantly reduced 60 days after therapy with AN-238. AN-238 also caused complete regression of MX-1 tumors in 5 of 10 animals, which remained tumor-free 60 days after treatment. In contrast, after treatment with cytotoxic radical AN-201, MDA-MB-231 and MCF-7-MIII tumors grew steadily and the regression of MX-1 tumors was only transitory in most animals. Toxicity of AN-201 was much greater than that of AN-238, as measured by animal deaths, loss of body weight and leukopenia. High-affinity sst receptors and mRNA for both sst(2) and sst(5) subtypes were found in all 3 tumor lines. Expression of sst receptors was not significantly affected by treatment with AN-238. Our results indicate that the cytotoxic somatostatin analog AN-238 efficaciously inhibits growth of human breast cancers expressing sst receptor subtypes 2 and 5.     

Somatostatin and dopamine receptors

The discovery of the new properties of SSRs and DRs has led to a renewed interest in agents targeting these receptors and has opened new perspectives for medical treatment of patients with pituitary and neuroendocrine tumors resistant to the "classical", currently available analogs. Moreover, SSRs and DRs crosstalk at membrane level may trigger alternative intracellular pathways or enhance the signalling for the control of cell growth. New somatostatin analogs and hybrid molecules, which display a broader and different spectrum of activities compared to conventional analogs, seem to be a promising therapeutic alternative for the control of hormone secretion and, hopefully, to reduce tumor burden. Receptor profile characterization is crucial for the accurate selection of patients potentially responsive to a given therapy.     

In vivo target modulation and biological activity of CHIR-258, a multitargeted growth factor receptor kinase inhibitor, in colon cancer models.

PURPOSE: To evaluate the therapeutic and biological effects of CHIR-258, an orally bioavailable, potent inhibitor of class III-V receptor tyrosine kinases, in colon cancer models. EXPERIMENTAL DESIGN: The pharmacologic activity of CHIR-258 was characterized by monitoring target modulation as well as by evaluating the antitumor and antiangiogenic effects in human colon xenograft models. RESULTS: CHIR-258 inhibits vascular endothelial growth factor receptor 1/2, fibroblast growth factor receptor 1/3, and platelet-derived growth factor receptor beta (PDGFRbeta) and shows both antitumor and antiangiogenic activities in vivo. Treatment of KM12L4a human colon cancer cells with CHIR-258 resulted in a dose-dependent inhibition of vascular endothelial growth factor receptor 1 and PDGFRbeta phosphorylation and reduction of phosphorylated extracellular signal-regulated kinase (ERK) levels, indicating modulation of target receptors and downstream signaling. In vivo administration of CHIR-258 resulted in significant tumor growth inhibition and tumor regressions, including large, established tumors (500-1,000 mm(3)). Immunohistochemical analysis showed a reduction of phosphorylated PDGFRbeta and phosphorylated ERK in tumor cells after oral dosing with CHIR-258 compared with control tumors. These changes were accompanied by decreased tumor cell proliferation rate and reduced intratumoral microvessel density. CHIR-258 inhibited the phosphorylation of PDGFRbeta and ERK phosphorylation in tumors within 2 hours following dosing and the inhibitory activity was sustained for >24 hours. Significant antitumor activity was observed with intermittent dosing schedules, indicating a sustained biological activity. CONCLUSION: These studies provide evidence that biological activity of CHIR-258 in tumors correlates with efficacy and aids in the identification of potential biomarkers of this multitargeted receptor tyrosine kinase inhibitor. CHIR-258 exhibits properties that make it a promising candidate for clinical development in a variety of solid and hematologic malignancies.     

The somatostatin analog Sandostatin (SMS201-995) in treatment of DMBA-induced rat mammary tumors

Summary The effects of treatment with a somatostatin analog (Sandostatin, SMS201-995) were investigated in female rats with dimethylbenzanthracene(DMBA)-induced rat mammary tumors. A 3-week treatment was performed using sandostatin, the LHRH-agonist buserelin alone, or buserelin in combination with sandostatin. Twice daily sandostatin treatment was performed with dosages of 0.05 µg, 0.2 µg, 1 µg, 5 µg, and 20 µg. Buserelin was used in a 2 × 5 µg/day dosage.The combined results from six different experiments show that the various dosages of sandostatin caused no tumor growth inhibition. Somatostatin receptors could not be demonstrated in these mammary tumors. Sandostatin treatment by daily injections did not suppress levels of growth hormone, prolactin, or epidermal growth factor-like activities. Estrogen (ER) and progesterone (PgR) receptor contents of the mammary tumors were not changed. In contrast, buserelin treatment caused highly significant tumor remission. The combined treatment with sandostatin and buserelin did not alter the treatment results obtained after treatment with buserelin alone.In conclusion, sandostatin treatment in this tumor model had no direct growth inhibitory effect and did not cause an endocrine inhibition of mammary tumor growth. However, these results do not exclude antitumor effects in human breast cancer in view of the presence of somatostatin receptors in approximately 20–45% of human tumors, besides possible different endocrine effects.     

Somatostatin inhibits the production of vascular endothelial growth factor in human glioma cells

In various cell types, the neuro- and endocrine peptide somatostatin induces inhibitory and anti-secretory effects. Since somatostatin receptors, especially of the subtype sst2A, are constantly over-expressed in gliomas, we investigated the influence of somatostatin and the receptor subtype-selective peptide/non-peptide agonists octreotide and L-054,522 on the secretion of the most important angiogenesis factor produced by gliomas, vascular endothelial growth factor (VEGF). Cultivated cells from solid human gliomas of different stages and glioma cell lines secreted variable amounts of VEGF, which could be lowered to 25% to 80% by co-incubation with somatostatin or sst2-selective agonists (octreotide and L-054,522). These effects were dose-dependent at nanomolar concentrations. Stimulation with different growth factors (EGF, bFGF) or hypoxia considerably increased VEGF production over basal levels. Growth factor-induced VEGF synthesis could be suppressed to <50% by co-incubation with somatostatin or an sst2-selective agonist; this was less pronounced in hypoxia-induced VEGF synthesis. The effects were detected at the protein and mRNA levels. These experiments indicate a potent anti-secretory action of somatostatin or sst2 agonists on human glioma cells that may be useful for inhibiting angiogenesis in these tumors.     

Gastrointestinal neuroendocrine tumors (NETs): new diagnostic and therapeutic challenges

This paper summarizes the current understanding of the biology of somatostatin receptor (sst), role of immunotherapy in neuroendocrine tumor (NET), new agents for PPRT, and methods to assess response and clinical benefit in NET. One of the most interesting aspects of sst biology is the recent discovery of truncated variants of the sst5 receptor subtype with unique tissue distribution and response to somatostatin (SST). These truncated receptors are associated with bad patient prognosis, decreased response to SST analogs, and may be new targets for diagnoses and treatment. IFN remains a cost-effective agent, particularly in classic mid gut carcinoids, and there is interest to continue examining immunotherapy's in this disease. PRRT remains a key strategy for treatment and imaging. In addition to the classic agents, there are a series of new agents targeting other receptors such as the incretin receptors (GLP-1R; GIPR) and other G-protein coupled receptors with great potential. With regards to therapy monitoring, the most commonly used criteria are Response Criteria Evaluation in Solid Tumors (RECIST). However, for different reasons, these criteria are not very useful in NET. Incorporation of other criteria such as Choi as well as functional imaging assessment with PET would be of great interest in this area.     

Diagnosis and treatment of carcinoid tumors

Carcinoid tumors belong to the family of neuroendocrine tumors, which are usually slow growing with distinct biological and clinical characteristics. The incidence of these tumors is approximately 2.5 in 100,000 people per year. The former classification system of foregut, midgut and hindgut tumors is still used in clinical routine, although there is a new World Health Organization classification. Determination of the histopathology of carcinoid tumors is of utmost importance and involves specific immunohistochemical staining for chromogranin A, synaptophysin, serotonin and gastrin. Proliferation capacity measured by Ki67 is used to guide forthcoming medical treatment. Localization procedures include computerized tomography, ultrasound, magnetic resonance imaging, somatostatin receptor scintigraphy and positron emission tomography. Surgery remains the cornerstone of treatment and provides the only chance of a cure. Other cytoreductive procedures include radiofrequency ablation, laser treatment and chemoembolization. Biological treatment includes cytotoxic agents, such as somatostatin analogs and interferon-alpha, which should be applied in slow-growing neoplasms. Combination regimens including cisplatin, etoposide, streptozotocin and 5-fluorouracil should be reserved for treatment of highly proliferating tumors. Future therapy of carcinoid tumors will be based on the specific tumor biology and treatment will be customized for each individual patient. New therapies, such as antiangiogenic agents and new, long-acting somatostatin analogs, together with further development of tumor-targeted treatments, will come into clinical use in the near future.     

Diagnosis and treatment of carcinoid tumors

Carcinoid tumors belong to the family of neuroendocrine tumors, which are usually slow growing with distinct biological and clinical characteristics. The incidence of these tumors is approximately 2.5 in 100,000 people per year. The former classification system of foregut, midgut and hindgut tumors is still used in clinical routine, although there is a new World Health Organization classification. Determination of the histopathology of carcinoid tumors is of utmost importance and involves specific immunohistochemical staining for chromogranin A, synaptophysin, serotonin and gastrin. Proliferation capacity measured by Ki67 is used to guide forthcoming medical treatment. Localization procedures include computerized tomography, ultrasound, magnetic resonance imaging, somatostatin receptor scintigraphy and positron emission tomography. Surgery remains the cornerstone of treatment and provides the only chance of a cure. Other cytoreductive procedures include radiofrequency ablation, laser treatment and chemoembolization. Biological treatment includes cytotoxic agents, such as somatostatin analogs and interferon-α, which should be applied in slow-growing neoplasms. Combination regimens including cisplatin, etoposide, streptozotocin and 5-fluorouracil should be reserved for treatment of highly proliferating tumors. Future therapy of carcinoid tumors will be based on the specific tumor biology and treatment will be customized for each individual patient. New therapies, such as antiangiogenic agents and new, long-acting somatostatin analogs, together with further development of tumor-targeted treatments, will come into clinical use in the near future.     

Somatostatin and cancer: applying endocrinology to oncology

Somatostatin is a peptide hormone that normally suppresses growth hormone (GH), thyrotropin (TSH), insulin and gut hormone release, as well as affecting multiple aspects of gastrointestinal function. It achieves these pleiotropic effects by binding somatostatin receptors (SSTR), a family of five G-protein coupled membrane receptors. Somatostatin analogs, such as octreotide, lanreotide and vapreotide, are well-established treatments for tumors that over secrete these hormones. Recently, use of somatostatin analogs for treating nonendocrine malignancies are being explored. Hu et al. found progressive reduction in SSTR3 expression when comparing normal gastric mucosa versus well differentiated versus poorly differentiated gastric adenocarcinomas; octreotide inhibited growth and induced apoptosis in vitro of those cells expressing SSTR3. Potential mechanisms by which somatostatin analogs may be useful in oncology include its endocrine actions, autocrine/paracrine effects, SSTR-mediated cell signaling and SSTR-mediated cell labeling.     

177Lu-DOTA(0),Tyr3] octreotate for somatostatin receptor-targeted radionuclide therapy

Receptor-targeted scintigraphy using radiolabeled somatostatin analogs such as octreotate is being used with great success to demonstrate the in vivo presence of somatostatin receptors on various tumors. A new and promising application for these analogs is radionuclide therapy. Radionuclides suitable for this application include the Auger electron-emitter (111)In and the beta-emitters (90)Y (high energy) and (177)Lu (low energy). We investigated [DOTA(0),Tyr(3)]octreotate, labeled with the lanthanide (177)Lu, in biodistribution and radionuclide therapy experiments using male Lewis rats bearing the somatostatin receptor-positive rat CA20948 pancreatic tumor. Biodistribution studies in Lewis rats showed the highest uptake in the rat pancreatic CA20948 tumor and sst(2)-positive organs, which include the adrenals, pituitary and pancreas, of [(177)Lu-DOTA(0),Tyr(3)]octreotate in comparison with (88)Y- and (111)In-labeled analogs. Kidney uptake of [(177)Lu-DOTA(0),Tyr(3)]octreotate could be reduced by approximately 40% by co-injection of 400 mg/kg D-lysine. In radionuclide therapy studies, a 100% cure rate was achieved in the groups of rats bearing small (< or =1 cm(2)) CA20948 tumors after 2 doses of 277.5 MBq or after a single dose of 555 MBq [(177)Lu-DOTA(0),Tyr(3)]octreotate. A cure rate of 75% was achieved after a single administration of 277.5 MBq. In rats bearing larger (> or =1 cm(2)) tumors, 40% and 50% cure rates were achieved in the groups that received 1 or 2 277.5 MBq injections of [(177)Lu-DOTA(0),Tyr(3)]octreotate, respectively. After therapy with [(177)Lu-DOTA(0),Tyr(3)]octreotide in rats bearing small tumors, these data were 40% cure after 1 injection with 277.5 MBq and 60% cure after 2 repeated injections. In conclusion, [(177)Lu-DOTA(0),Tyr(3)]octreotate has demonstrated excellent results in radionuclide therapy studies in rats, especially in animals bearing smaller tumors. This candidate molecule shows great promise for radionuclide therapy in patients with sst(2)-expressing tumors.     

Somatostatin and somatostatin analogues: diagnostic and therapeutic uses.

Somatostatin and its octapeptide analogues exert their effects through interaction with somatostatin receptor (sst) subtypes 1 through 5 (sst 1-5 ). Somatostatin binds with high affinity to all sst subtypes, whereas the currently commercially available octapeptide analogues bind only with a high affinity to sst 2 and sst 5. Pituitary tumors, endocrine pancreatic tumors, and carcinoid tumors express multiple sst subtypes, but sst 2 predominance is found in 90% of carcinoids and 80% of endocrine pancreatic tumors. Sst 2 and sst 5 predominance is found in growth hormone-secreting pituitary tumors. In patients harboring sst 2 - or sst 5 -positive neuroendocrine tumors, clinical symptomatology can be controlled by the chronic administration of one of the currently commercially available octapeptide somatostatin analogues. Tumors and metastases that bear sst 2 or sst 5 can be visualized in vivo after injection of radiolabeled octapeptide analogues. Radiolabeled octapeptide analogues can also be used for radiotherapy of sst 2 - and sst 5 -positive advanced or metastatic neuroendocrine tumors.     

Targeted therapy with a cytotoxic somatostatin analog, AN-238, inhibits growth of human experimental endometrial carcinomas expressing multidrug resistance protein MDR-1

BACKGROUND: Chemoresistance mediated by membrane transporters such as multidrug resistance (MDR-1) glycoprotein remains a challenge in the chemotherapy treatment of advanced or recurrent endometrial carcinoma. Targeted chemotherapy might overcome this resistance. The cytotoxic somatostatin (SST) analog, AN-238, consists of a superactive derivative of doxorubicin (DOX), 2-pyrrolino-DOX (AN-201), linked to the SST analog carrier, RC-121. This conjugate binds strongly to SST receptor subtypes (sst) 2a (sst2(a)) and 5 (sst(5)) and can be targeted to tumors that express these receptors. METHODS: The presence of sst2(a) and sst(5) was determined in 3 human endometrial carcinoma cell lines (HEC-1A, RL-95-2, and AN3CA). Nude mice bearing xenografts of these cancers were treated with AN-238 and its radical, AN-201. The antitumor effects and toxicity were compared. The authors studied the effects of AN-238 and AN-201 on the expression levels of MDR-1, multidrug resistance related protein (MRP-1), and breast carcinoma resistance protein (BCRP) by real-time polymerase chain reaction. RESULTS: The authors demonstrated the presence of mRNA and receptor protein for sst(2a) and sst(5) on HEC-1A, RL-95-2, and AN3CA tumors. AN-238 significantly (P < 0.05) inhibited the growth of these tumors, whereas AN-201 had no effect. Blockade of SST receptors nullified the effects of AN-238. In all 3 endometrial carcinoma lines, AN-238 caused a weaker induction of MDR-1 than AN-201. No major induction of MRP-1 and BCRP occurred after treatment with AN-238 or AN-201. CONCLUSIONS: Targeted chemotherapy with the cytotoxic SST analog, AN-238, inhibited powerfully the growth of endometrial carcinoma, which express SST receptors, regardless of their expression level of MDR-1.