Clinical application of Fluciclovine PET, choline PET and gastrin-releasing polypeptide receptor (bombesin) targeting PET in prostate cancer.

PURPOSE OF REVIEW: The aim of this review is to explore the clinical application of different PET radiopharmaceuticals in prostate cancer (PCa), beyond inhibitors of the prostate-specific membrane antigen (PSMA). RECENT FINDINGS: Choline PET represented in the last decades the standard of reference for PET imaging in PCa and has been recently included in clinical trials evaluating the efficacy of metastasis-directed therapy in oligo-metastatic disease. Fluciclovine, as synthetic amino acid, has been proposed for investigating PCa. The results obtained by the first prospective studies led to FDA approval in 2016 in patients with biochemical recurrence. Recently, phase II/III trials explored its accuracy compared with PSMA PET and its impact on patient management. Imaging the gastrin-releasing polypeptide receptor (GRPR) recently drawn attention. Radio-labelled GRPR antagonists have the potential to be used as theranostic agents. Further evaluation is needed to understand the relation between GRPR expression and hormonal-resistant PCa, and for tumors characterized by heterogeneity of receptors expressed (e.g. PSMA-negative) on their cell surface. SUMMARY: Other new generation PET tracers may play an important role in PCa, namely in case of PSMA-negative phenotypes.

Growth factors and medium hyperglycemia induce Sox9+ ductal cell differentiation into ß cells in mice with reversal of diabetes.

We previously reported that long-term administration of a low dose of gastrin and epidermal growth factor (GE) augments ß-cell neogenesis in late-stage diabetic autoimmune mice after eliminating insulitis by induction of mixed chimerism. However, the source of ß-cell neogenesis is still unknown. SRY (sex-determining region Y)-box 9(+) (Sox9(+)) ductal cells in the adult pancreas are clonogenic and can give rise to insulin-producing ß cells in an in vitro culture. Whether Sox9(+) ductal cells in the adult pancreas can give rise to ß cells in vivo remains controversial. Here, using lineage-tracing with genetic labeling of Insulin- or Sox9-expressing cells, we show that hyperglycemia (>300 mg/dL) is required for inducing Sox9(+) ductal cell differentiation into insulin-producing ß cells, and medium hyperglycemia (300-450 mg/dL) in combination with long-term administration of low-dose GE synergistically augments differentiation and is associated with normalization of blood glucose in nonautoimmune diabetic C57BL/6 mice. Short-term administration of high-dose GE cannot augment differentiation, although it can augment preexisting ß-cell replication. These results indicate that medium hyperglycemia combined with long-term administration of low-dose GE represents one way to induce Sox9(+) ductal cell differentiation into ß cells in adult mice.

Indian Hedgehog mediates gastrin-induced proliferation in stomach of adult mice.

BACKGROUND & AIMS: Loss of expression of Sonic Hedgehog (Shh) from parietal cells results in hypergastrinemia in mice, accompanied by increased expression of Indian Hedgehog (Ihh) and hyperproliferation of surface mucous cells. We investigated whether hypergastrinemia induces gastric epithelial proliferation by activating Ihh signaling in mice. METHODS: We studied mice with parietal cell-specific deletion of Shh (PC-Shh(KO)) and hypergastrinemia, crossed with gastrin-deficient (GKO) mice (PC-Shh(KO)/GKO). When mice were 3-4 months old, gastric tissues were collected and analyzed by histology, for incorporation of bromodeoxyuridine, and for expression of the surface mucous cell marker Ulex europaeus. PC-Shh(KO)/GKO mice were given gastrin infusions for 7 days; gastric surface epithelium was collected and expression of Ihh was quantified by laser capture microdissection followed by quantitative reverse transcriptase polymerase chain reaction. Mouse stomach-derived organoids were incubated with or without inhibitors of WNT (DKK1) or Smoothened (vismodegib) and then cocultured with immortalized stomach mesenchymal cells, to assess proliferative responses to gastrin. RESULTS: Gastric tissues from PC-Shh(KO)/GKO mice with hypergastrinemia had an expanded surface pit epithelium, indicated by a significant increase in numbers of bromodeoxyuridine- and Ulex europaeus-positive cells, but there was no evidence for hyperproliferation. Gastrin infusion of PC PC-Shh(KO)/GKO mice increased expression of Ihh and proliferation within the surface epithelium compared with mice given infusions of saline. In gastric organoids cocultured with immortalized stomach mesenchymal cells, antagonists of WNT and Smoothened inhibited gastrin-induced proliferation and WNT activity. Activity of WNT in media collected from immortalized stomach mesenchymal cells correlated with increased expression of glioma-associated oncogene homolog 1, and was inhibited by DKK1 or vismodegib. CONCLUSIONS: Ihh signaling mediates gastrin-induced proliferation of epithelial cells in stomachs of adult mice.

Omeprazole and PGC-formulated heparin binding epidermal growth factor normalizes fasting blood glucose and suppresses insulitis in multiple low dose streptozotocin diabetes model.

PURPOSE: Our objective was to develop novel nanocarriers (protected graft copolymer, PGC) that improve the stability of heparin binding EGF (HBEGF) and gastrin and then to use PGC-formulated HBEGF (PGC-HBEGF) and Omeprazole (+/- PGC-gastrin) for normalizing fasting blood glucose (FBG) and improving islet function in diabetic mice. METHODS: HBEGF, PGC-HBEGF, Omeprazole, Omeprazole + PGC-HBEGF, Omeprazole + PGC-gastrin + PGC-HBEGF and epidermal growth factor (EGF) + gastrin were tested in multiple low dose streptozotocin diabetic mice. RESULTS: Omeprazole + PGC-HBEGF normalized FBG and is better than EGF + gastrin at improving islet function and decreasing insulitis. Groups treated with Omeprazole, Omeprazole + PGC-HBEGF, or EGF + gastrin have significantly improved islet function versus saline control. All animals that received PGC-HBEGF had significantly reduced islet insulitis versus saline control. Non-FBG was lower for Omeprazole + PGC-gastrin + PGC-HBEGF but Omeprazole + PGC-HBEGF alone showed better FBG and glucose tolerance. CONCLUSIONS: Omeprazole + PGC-HBEGF provides a sustained exposure to both EGFRA and gastrin, improves islet function, and decreases insulitis in multiple low dose streptozotocin diabetic mice. Although HBEGF or EGF elevates non-FBG, it facilitates a reduction of insulitis and, in the presence of Omeprazole, provides normalization of FBG at the end of treatment. The study demonstrates Omeprazole and PGC-HBEGF is a viable treatment for diabetes.

Gastric type I carcinoid: a pilot study with human G17DT immunogen vaccination.

CONTEXT: Gastric type I carcinoid is a rare neoplasm, deriving from enterochromaffin-like cells (ECL), mainly affecting women with autoimmune gastritis. The approach to treatment, either endoscopic, medical or surgical, is not well defined, particularly in multifocal tumours or carcinoids with rapid growth/frequent recurrence. OBJECTIVE: To determine whether an anti-G17 vaccination might interfere on the natural history of gastric type I carcinoid. SETTING: Padua teaching Hospital, outpatient clinic. DESIGN AND PATIENTS: Three patients with type I gastric carcinoid in autoimmune gastritis were administered, after informed consent and ethic committee approval, with a vaccine against gastrin 17 (G17), a synthetic peptide that stimulates specific and high-affinity anti-G17 antibodies, and followed up endoscopically and clinically for a mean of 36 months. MAIN OUTCOME MEASURES: Gastric histology and specifically carcinoid growth/recurrence and trend in time in gastrin, G17, pepsinogens, chromogranin A and clinical parameters. RESULTS: Following vaccination, carcinoid regression was observed in 2/3 patients and, in one of the patients, even the disappearance of ECL hyperplasia, with a reduced ECL cells stimulation, confirmed by a significant reduction in chromogranin A levels. Regression was observed in the two patients that showed a more clear local response to the vaccine. Increased autoantibody titre was observed, but no appearance of new autoimmune diseases. CONCLUSIONS: Anti-G17 vaccination induced regression of type I gastric carcinoid and could be considered for the treatment of this tumour, when endoscopic removal is not indicated.

Stimulation of proliferation in the colorectal mucosa by gastrin precursors is blocked by desferrioxamine.

Precursors of the peptide hormone gastrin stimulate proliferation in the colorectal mucosa and promote the development of colorectal carcinoma. Gastrins bind two ferric ions selectively and with high affinity, and the biological activity of glycine-extended gastrin (Ggly) in vitro is dependent on the presence of ferric ions. The aim of the present study was to determine whether or not iron is required for biological activity of progastrin and Ggly in vivo. Rats that had undergone a colostomy were infused with Ggly, and proliferation was measured in the defunctioned rectal mucosa. Proliferation was also measured in the colonic mucosa of hGAS and MTI-Ggly mice, which, by definition, overexpress progastrin and Ggly, respectively. The requirement for iron was assessed by thrice-weekly injection of the chelating agent desferrioxamine (DFO). The proliferation index in the defunctioned rectal mucosa was significantly increased in the Ggly-infused rats, and the increase was significantly reduced after treatment with DFO. Treatment with DFO significantly reduced the crypt height and proliferation index in the colonic mucosa of hGAS and MTI-Ggly mice but had no effect on the same variables in wild-type mice. These observations are consistent with the hypothesis that the biological activity of progastrin and Ggly in vivo is dependent on the presence of ferric ions and further suggest that chelating agents may block the stimulatory effects of gastrin precursors in the development of colorectal carcinoma.

Antidiabetic effects and sustained metabolic benefits of sub-chronic co-administration of exendin-4/gastrin and xenin-8-Gln in high fat fed mice.

The present study has examined the antidiabetic effects of 21 days co-administration of xenin-8-Gln with the dual-acting fusion peptide, exendin-4/gastrin, as well as persistence of beneficial metabolic benefits, in high fat fed (HFF) mice. Xenin-8-Gln, exendin-4 and gastrin represent compounds that activate receptors of the gut-derived hormones, xenin, glucagon-like peptide-1 (GLP-1) and gastrin, respectively. Twice-daily administration of exendin-4/gastrin, xenin-8-Gln or a combination of both peptides significantly reduced circulating glucose, HbA1c and cumulative energy intake. Combination therapy with xenin-8-Gln and exendin-4/gastrin increased circulating insulin. All HFF mice treated with exendin-4/gastrin presented with body weight similar to lean control mice on day 21. Each treatment improved glucose tolerance and the glucose-lowering actions of glucose dependent insulinotropic polypeptide (GIP), as well as augmenting glucose- and GIP-induced insulin secretion, with benefits being most prominent in the combination group. Administration of exendin-4/gastrin alone, and in combination with xenin-8-Gln, increased pancreatic insulin content and improved the insulin sensitivity index. Pancreatic beta-cell area was significantly increased, and alpha cell area decreased, by all treatments, with the combination group also displaying enhanced overall islet area. Notably, metabolic benefits were generally retained in all groups of HFF mice, and especially in the combination group, following discontinuation of the treatment regimens for 21 days. This was associated with maintenance of increased islet and beta-cell areas. Together, these data confirm the antidiabetic effects of co-activation of GLP-1, gastrin and xenin cell signalling pathways, and highlight the sustainable benefits this type of treatment paradigm can offer in T2DM.

Synergistic action of gastrin and ghrelin on gastric acid secretion in rats.

Gastrin and ghrelin are secreted from G cells and X/A-like cells in the stomach, respectively, and respective hormones stimulate gastric acid secretion by acting through histamine and the vagus nerve. In this study, we examined the relationship between gastrin, ghrelin and gastric acid secretion in rats. Intravenous (iv) administration of 3 and 10 nmol of gastrin induced transient increases of ghrelin levels within 10 min in a dose-dependent manner. Double immunostaining for ghrelin and gastrin receptor revealed that a proportion of ghrelin cells possess gastrin receptors. Although (iv) administration of gastrin or ghrelin induced significant gastric acid secretion, simultaneous treatment with both hormones resulted in a synergistic, rather than additive, increase of gastric acid secretion. This synergistic increase was not observed in vagotomized rats. These results suggest that gastrin may directly stimulate ghrelin release from the stomach, and that both hormones may increase gastric acid secretion synergistically.

Targeted Magnetic Intra-Lysosomal Hyperthermia produces lysosomal reactive oxygen species and causes Caspase-1 dependent cell death.

Therapeutic strategies using drugs which cause Lysosomal Cell Death have been proposed for eradication of resistant cancer cells. In this context, nanotherapy based on Magnetic Intra-Lysosomal Hyperthermia (MILH) generated by magnetic nanoparticles (MNPs) that are grafted with ligands of receptors overexpressed in tumors appears to be a very promising therapeutic option. However, mechanisms whereby MILH induces cell death are still elusive. Herein, using Gastrin-grafted MNPs specifically delivered to lysosomes of tumor cells from different cancers, we provide evidences that MILH causes cell death through a non-apoptotic signaling pathway. The mechanism of cell death involves a local temperature elevation at the nanoparticle periphery which enhances the production of reactive oxygen species through the lysosomal Fenton reaction. Subsequently, MILH induces lipid peroxidation, lysosomal membrane permeabilization and leakage of lysosomal enzymes into the cytosol, including Cathepsin-B which activates Caspase-1 but not apoptotic Caspase-3. These data highlight the clear potential of MILH for the eradication of tumors overexpressing receptors.

Pure human big gastrin. Immunochemical properties, disappearance half time, and acid-stimulating action in dogs.

Biological properties of pure natural human "big gastrin" (designated G-34 because it contains 34 amino acid residues) were compared with those of pure natural heptadecapeptide gastrins (G-17) from human and porcine sources. Radioimmunoassay inhibition curves indicated that G-17 was nearly 1.5 times more potent than G-34 with the antibody used in this study. This difference was confirmed by demonstration of increased immunoreactivity generated when G-34 was converted to G-17 by trypsinization. When infused intravenously into dogs with gastric fistulas and Heidenhain pouches in equimolar doses, G-34 produced slightly higher acid secretory responses than G-17. Responses to sulfated and nonsulfated forms were not significantly different, nor were responses to human and porcine G-17. During infusion of equimolar doses, steady-state serum gastrin concentrations were more than fivefold higher with G-34 than with G-17. The difference in steady-state blood concentrations could be accounted for by a corresponding difference in removal rates. The half times of the G-34 preparations averaged 15.8 min and the half times of the G-17 preparations averaged 3.2 min. The calculated spaces of distribution for G-17 and G-34 were similar, about 25% of body weight. When the increment in serum gastrin was plotted against acid secretory response it was found that nearly five times greater increments in molar concentrations of G-34 than of G-17 were required to produce the same rate of acid secretion. The potency of these two molecular forms of gastrin can be expressed in two different ways. Based on exogenous molar doses, the potencies of G-34 and G-17 were similar. However, based on molar increments in serum gastrin concentration, G-17 was approximately five times more potent than G-34. Hence, fractionation of these gastrin components may be important in estimation of the acid-stimulating action represented by total serum gastrin as measured by radio-immunoassay.

The effect of gastrin on basal- and glucose-stimulated insulin secretion in man.

The effect of gastrin on basal- and glucose-stimulated insulin secretion was studied in 32 normal, young subjects. The concentration of gastrin and insulin in serum was measured radioimmunochemically. Maximal physiologic limit for the concentration of gastrin in serum was of the order of 160 pmol per liter as observed during a protein-rich meal. Oral ingestion of 50 g glucose produced a small gastrin response from 28+/-3 to 39+/-5 pmol per liter (mean +/-SEM, P < 0.01). Intravenous injection or prolonged infusion of gastrin increased the concentration of insulin in peripheral venous blood to a maximum within 2 min followed by a decline to basal levels after a further 10 min. The minimum dose required to induce a significant insulin response (31.2 ng gastrin per kg) increased the gastrin level in serum above the physiologic range. Maximum effect was obtained with 500 ng gastrin per kg. When 15.6 ng (7.1 pmol) gastrin per kg body weight and 25 g glucose were injected simultaneously, the glucose-induced insulin response was potentiated (from 2.32+/-0.33 to 4.33+/-0.98 nmol per liter per 20 min, P < 0.02), even though gastrin concentrations only increased to 71.2+/-6.6 pmol per liter. No effect, however, was noted on glucose disposal. 15.6 ng gastrin per kg given i.v. 30 min before an i.v. glucose tolerance test was without significant effect on the insulin response. The results indicate that gastrin can stimulate a rapid and short-lived release of insulin. In physiologic concentrations gastrin potentiates the glucose-stimulated insulin secretion and is without effect on basal insulin secretion. A small release of gastrin during oral glucose ingestion may to a limited extent contribute to the nonglycemic insulin secretion. During protein ingestion, gastrin probably stimulates insulin secretion significantly.

Z-360 Suppresses Tumor Growth in MIA PaCa-2-bearing Mice via Inhibition of Gastrin-induced Anti-Apoptotic Effects.

BACKGROUND/AIM: The aim of the study was to evaluate the anti-tumor mechanism of Z-360, a gastrin/cholecystokinin-2 receptor (CCK2R) antagonist, in MIA PaCa-2 cells and in a subcutaneous xenograft mice model. MATERIALS AND METHODS: The anti-tumor effects of Z-360 and/or gemcitabine were monitored using a MIA PaCa-2 xenograft model. The effect of Z-360 on apoptosis in the model was examined by TUNEL staining and real-time PCR analysis and the effect in MIA PaCa-2 cells stably expressing human CCK2R was also evaluated by caspase-3/7 activity. RESULTS: In this xenograft model, Z-360 significantly reduced the tumor weight, increased TUNEL-positive cells and suppressed the expression of anti-apoptosis factors such as survivin, XIAP and Mcl-1, and these effects of Z-360 combined with gemcitabine were more effective. Furthermore, gastrin-17 and gastrin-34 inhibited apoptosis in vitro and Z-360 dose-dependently abrogated this effect. CONCLUSION: These results suggest that Z-360 exerts an anti-tumor effect through a reduction in anti-apoptosis factors by blocking CCK2R.

Combination therapy with epidermal growth factor and gastrin increases beta-cell mass and reverses hyperglycemia in diabetic NOD mice.

Combination therapy with epidermal growth factor (EGF) and gastrin induces beta-cell regeneration in rodents with chemically induced diabetes. We investigated whether EGF plus gastrin could correct hyperglycemia in NOD mice with autoimmune diabetes. Combined treatment with EGF (1 mug/kg) and gastrin (3 mug/kg) for 2 weeks restored normoglycemia after diabetes onset in NOD mice, whereas EGF or gastrin alone did not. Fasting blood glucose remained normal (3.5-6.5 mmol/l) or mildly elevated (<11 mmol/l) in five of six mice (83%) for 10 weeks after EGF plus gastrin treatment was stopped, whereas all mice treated with vehicle or EGF or gastrin alone became severely hyperglycemic (12-35 mmol/l). Pancreatic beta-cell mass was increased threefold and insulin content was increased eightfold in mice treated with EGF plus gastrin compared with pretreatment values. The correction of hyperglycemia correlated significantly with increases in pancreatic beta-cell mass and insulin content. In addition, splenic cells from mice treated with EGF plus gastrin delayed diabetes induction by adoptive transfer of diabetogenic cells into immunodeficient NOD-scid mice, suggesting the induction of immunoregulatory cells in NOD mice treated with EGF plus gastrin. We conclude that a short course of combined EGF and gastrin therapy increases pancreatic beta-cell mass and reverses hyperglycemia in acutely diabetic NOD mice; the impact of this combined therapy may result from the effects of EGF and gastrin on beta-cells, immune cells, or both.

Gastrin protection against chemically induced gastric adenocarcinomas in Wistar rats: histopathology of the glandular stomach and incidence of gastric adenocarcinoma.

The effects of gastrin on the histopathology of the glandular stomach and on the incidence of gastric carcinoma induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were investigated in inbred Wistar (W) rats. Prolonged administration of gastrin after treatment with MNNG significantly reduced the incidence of adenocarcinomas of the glandular stomach. In addition, atypical glandular proliferations were significantly less frequent and were smaller, and the incidence of marked mucosal atrophy was significantly reduced in both the antral and oxyntic gland mucosae. Both atypical glandular hyperplasia and mucosal atrophy are precursors of gastric cancers; prolonged administration of gastrin to rats after treatment with MNNG suppressed development of precursors of gastric cancer and so prevented development of gastric cancers.

Effect of prolonged administration of gastrin on experimental carcinogenesis in rat colon induced by intrarectal instillation of N-methyl-N'-nitro-N-nitrosoguanidine.

The effect of tetragastrin on the incidence and histology of colonic tumors induced by intrarectal instillation of N-methyl-N'-nitro-N-nitrosoguanidine was investigated in Wistar rats. Prolonged administration of tetragastrin in depot form during and after treatment with N-methyl-N'-nitro-N-nitrosoguanidine resulted in a significant reduction in the incidence of colonic tumors in Experimental Week 35. Histological examinations showed that, unlike the well-differentiated adenocarcinomas with a typical glandular pattern in control groups, the adenocarcinomas that developed in rats treated with tetragastrin had high mucin-producing activity.

Hypergastrinemia promotes adenoma progression in the APC(Min-/+) mouse model of familial adenomatous polyposis.

Serum hypergastrinemia promotes the growth of colorectal adenocarcinoma. Some colorectal adenomas express cholecystokinin B/gastrin receptor mRNA, and thus hypergastrinemia may increase progression through the adenoma-carcinoma sequence. This was investigated in the multiple intestinal neoplasia APC(Min-/+) mouse. Serum gastrin levels in APC(Min-/+) mice were elevated 5-6-fold by oral administration of omeprazole (75 mg/kg). Terminal tumor burden was monitored by onset of anemia. A labeling index was generated by immunohistochemical detection of bromodeoxyuridine incorporation. Serum gastrin was neutralized by antigastrin antibodies raised in situ by use of a gastrin immunogen, Gastrimmune. Hypergastrinemia resulted in reduced survival of the APC(Min-/+) mice from a median survival of 13 weeks in the controls to 10 weeks following omeprazole treatment (P < 0.00001, log-rank test). The labeling indices of adenomas from the small and large intestines of omeprazole-treated mice were increased 35 and 29%, respectively (P < 0.05 and P < 0.025, respectively). Gastrimmune immunization reversed both the survival effect and the increased proliferation resulting from serum hypergastrinemia. Hypergastrinemia may promote the progression of existing premalignant colonic lesions by increasing proliferation. Clinical investigations should determine whether this occurs in the human scenario, considering the widespread use of proton pump inhibitors.

[The use of a new analogue DGlu-Octagastrin in scintigraphy of medullary thyroid carcinoma]. / Zastosowanie nowego analogu DGlu-Oktagastryny w scyntygrafii raka rdzeniatego tarczycy.

INTRODUCTION: Medullary thyroid carcinomas (MTC) reveal overexpression of several peptide receptors particularly of gastrin and cholecystokinin 2 (CCK2). Experimental studies of various CCK-2/gastrin analogues found that a C-terminal, 8-aminoacid peptide, (D)Glu-octagastrin, has optimal properties. Thus, the aim of our studies was to prepare (99m)Tc-labelled HYNIC-octagastrin, to evaluate its biologic tolerance in animals and introduce for scintigraphy in patients with MTC. MATERIAL AND METHODS: HYNIC-(D)Glu-octagastrin was from piCHEM (Graz, Austria), (99m)Tc generator from Amersham (Health), other reagents were purchased from Sigma. Labelling of the peptide was performed in phosphate buffer of pH 6.0 for 10 minutes at 100(o)C using EDDA and tricine as coligands. RESULTS: The labeling yields were high (above 95%); the specific activity amounted to 1200 to 1430 microCi/microg. Radiochemical purity on SepPak cartridge and ITLC ranged from 94 to 98%. No adverse effects were observed in mice after administration of 10 to 50 times greater doses that those used in patients. Clinical studies comprised 20 patients with MTC and high serum calcitonin. (99m)Tc-EDDA/HYNIC-(D)Glu-octagastrin, 500 to 700 MBq, was administered iv and whole body scintigraphy was performed using a double head gamma-camera (Varicam, Elscint) 2 and 4 hours later. Increased accumulation of the tracer in foci of MTC and its metastases was found in 8 patients. CONCLUSIONS: Scintigraphy with a new gastrin analogue ((D)Glu-octagastrin) makes it possible to detect MTC with overexpression of CCK-2/gastrin receptors and to select patients for receptor-mediated radiopeptide therapy using DOTA-gastrin analogues labelled with (177)Lu and (90)Y.

Reduced ghrelin, islet amyloid polypeptide, and peptide YY expression in the stomach of gastrin-cholecystokinin knockout mice.

The antral hormone gastrin and its intestinal relative, cholecystokinin (CCK), are pivotal in the regulation of gastric functions. Other gastric hormones like ghrelin, peptide YY (PYY), and islet amyloid polypeptide (IAPP), however, also contribute to the regulation of acid secretion, motility, and feeding. Because gastrin and CCK are crucial for gastric homeostasis, we examined how loss of gastrin alone and gastrin plus CCK affected the expression of ghrelin, IAPP, and PYY and ghrelin secretion. The expression of ghrelin, IAPP, and PYY and the CCK-A receptor genes were examined in both gastrin and gastrin-CCK double-knockout (KO) mice using immunocytochemistry and quantitative RT-PCR. Ghrelin concentrations in plasma were measured using RIA. Gastrin and CCK were infused in gastrin-CCK KO mice using osmotic minipumps. The number of ghrelin cells and ghrelin gene expression were unaffected, albeit the ghrelin cells were located closer to the base of the glands in both KO mouse strains when freely fed. However, lack of both gastrin and CCK attenuated fasting-induced ghrelin expression and secretion. Fundic ghrelin cells expressed the CCK-A receptor, and ghrelin expression increased after CCK infusion. Furthermore, gastric IAPP and PYY expression as well as the number of IAPP- and PYY-containing cells were reduced in both gastrin and gastrin-CCK KO mice. Gastrin infusion increased gastric IAPP but not PYY expression. In conclusion, lack of gastrin plus CCK but not gastrin alone reduced ghrelin secretion in response to fasting through both direct and indirect mechanisms. Both gastrin and combined gastrin-CCK deficiency reduced the gastric IAPP and PYY expression.

Radiolabeled peptides for targeting cholecystokinin-B/gastrin receptor-expressing tumors.

UNLABELLED: The high sensitivity of pentagastrin stimulation in detecting primary or metastatic medullary thyroid cancer (MTC) suggests widespread expression of the corresponding receptor type on human MTC. Indeed, autoradiographic studies have demonstrated cholecystokinin (CCK)-B/gastrin receptors not only in more than 90% of MTCs but also in a high percentage of small cell lung cancers, some ovarian cancers, astrocytomas and potentially a variety of adenocarcinomas. The aim of this study was to systematically screen and optimize, in a preclinical model and a pilot clinical study, suitable radioligands for targeting CCK-B receptors in vivo. METHODS: A variety of CCK/gastrin-related peptides, all bearing the C-terminal CCK receptor-binding tetrapeptide sequence Trp-Met-Asp-PheNH2 or derivatives thereof, were studied. They were radioiodinated by the lodogen or Bolton-Hunter procedures. The peptides were members of the gastrin or CCK families, which differ by the intramolecular position of a tyrosyl moiety. Their stability and affinity were studied in vitro and in vivo; their biodistribution and therapeutic efficacy were tested in nude mice bearing subcutaneous human MTC xenografts. Diethylenetriamine pentaacetic acid (DTPA) derivatives of suitable peptides were synthesized successfully, and their preclinical and initial clinical evaluations were performed, labeled with 111In. RESULTS: All members of the CCK or gastrin families were stable in serum (with half-lives of several hours at 37 degrees C); nevertheless, the stability of those peptides bearing N-terminal pGlu residues or D-amino acids was significantly higher. In accordance with their comparably low affinity, nonsulfated members of the CCK family showed fairly low uptake in the tumor and other CCK-B receptor-expressing tissues. Sulfated CCK derivatives performed significantly better but also displayed a comparably high uptake in normal CCK-A receptor-expressing tissues. This effect was probably due to their similar affinity for both CCK-A and CCK-B receptors. Best tumor uptake and tumor-to-nontumor ratios were obtained with members of the gastrin family because of their selectivity and affinity for the CCK-B receptor subtype. Pilot therapy experiments in MTC-bearing animals showed significant antitumor efficacy compared with untreated controls. DTPA derivatives of minigastrin were successfully developed. In a pilot clinical study, radioiodinated and 111In-labeled derivatives showed excellent targeting of physiological CCK-B receptor-expressing organs, as well as all known tumor sites. CONCLUSION: CCK/gastrin analogs may be a useful new class of receptor-binding peptides for diagnosis and therapy of CCK-B receptor-expressing tumors, such as MTC or small cell lung cancer. Nonsulfated gastrin derivatives may be preferable because of their CCK-B receptor selectivity, hence lower accretion in normal CCK-A receptor-expressing organs.

ECL-cell histamine mobilization in conscious rats: effects of locally applied regulatory peptides, candidate neurotransmitters and inflammatory mediators.

1. The ECL cells control gastric acid secretion by mobilizing histamine in response to circulating gastrin. In addition, the ECL cells are thought to operate under nervous control and to be influenced by local inflammatory processes. 2. The purpose of the present study was to monitor histamine mobilization from ECL cells in conscious rats in response to locally applied regulatory peptides, candidate neurotransmitters and inflammatory mediators. 3. Microdialysis probes were implanted in the submucosa of the acid-producing part of the rat stomach. Three days later, the agents to be tested were administered via the microdialysis probe and their effects on basal (48 h fast) and stimulated (intravenous infusion of gastrin-17, 3 nmol kg(-1) h(-1)) mobilization of ECL-cell histamine was monitored by continuous measurement of histamine in the perfusate (radioimmunoassay). 4. Locally administered gastrin-17 and sulfated cholecystokinin-8 mobilized histamine as did pituitary adenylate cyclase-activating peptide-27, vasoactive intestinal peptide, peptide YY, met-enkephalin, endothelin and noradrenaline, adrenaline and isoprenaline. 5. While gastrin, sulfated-cholecystokinin-8, met-enkephalin and isoprenaline induced a sustained elevation of the submucosal histamine concentration, endothelin, peptide YY, pituitary adenylate cyclase activating peptide, vasoactive intestinal peptide, noradrenaline and adrenaline induced a transient elevation. 6. Calcitonin gene-related peptide, galanin, somatostatin and the prostanoid misoprostol inhibited gastrin-stimulated histamine mobilization. 7. The gut hormones neurotensin and secretin and the neuropeptides gastrin-releasing peptide, neuropeptide Y and substance P failed to affect ECL-cell histamine mobilization, while motilin and neuromedin U-25 had weak stimulatory effects. Also acetylcholine, carbachol, serotonin and the amino acid neurotransmitters aspartate, gamma-aminobutyric acid, glutamate and glycine were inactive or weakly active as was bradykinin. 8. In summary, a range of circulating hormones, local hormones, catecholamines, neuropeptides and inflammatory mediators participate in controlling the activity of rat stomach ECL cells in situ.