The effect of somatostatin on the production of human interferons by mononuclear cells.

Somatostatin (SMS) is a tetradecapeptide which can inhibit the secretion of a number of peptides produced by the endocrine or nervous systems. SMS 201-995 (octreotide) is a somatostatin analogue with very potent somatostatin activities. We have been investigating the effects of both SMS and octreotide on the production of human interferon (IFN). We obtained human peripheral blood mononuclear cells from normal donors and induced them to produce IFN in the presence or absence of a number of peptides possessing somatostatin activities. SMS and octreotide were shown to inhibit the secretion of INF-gamma but not IFN-alpha. Concentrations of 10(-6) M were shown to decrease yields when Concanavalin A or phytohemagglutin were used as the inducer. Higher concentrations had a progressively greater effect. No effects were observed on IFN-gamma production if interleukin 2, ionomycin, or various natural antigens were used to induce the cells. The 28-amino acid form of somatostatin had some effects on gamma IFN yields but the first 14-amino acid fragment of this peptide moiety did not. No effect of any of these compounds was observed on IFN bioactivity. These studies indicate SMS may have some regulatory action on the secretion of immunomodulators in vitro but the concentrations required are well above those encountered under physiologic circumstances, suggesting SMS may not play an important regulatory role governing such secretion in vivo.     

The effect of a somatostatin analogue (SMS 201-995, Sandostatin) on the concentration of phosphoribosyl pyrophosphate and the activity of the pentose phosphate pathway in the early renal hypertrophy of experimental diabetes in the rat

The effect of a long-acting somatostatin analogue on the acute renal hypertrophy following induction of experimental diabetes in the rat has been studied. The kidney weight increase occurring at 2 and 7 days after alloxan injection was significantly lower in the diabetic group receiving somatostatin. Similarly, the previously reported increase in glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (EC 1.1.1.44) found in the kidney at 2 and 7 days of diabetes was less marked in the group receiving SMS 201-995. The fall in renal phosphoribosyl pyrophosphate associated with early diabetic renal hypertrophy (7) was also lessened by administration of SMS 201-995. No effects of the drug were found in the normal rat on the same regimen of treatment. These observations indicate involvement of glucagon and/or growth hormone in the initiation of kidney growth in diabetes.     

Suppression of growth hormone and somatomedin C by long-acting somatostatin analog SMS 201-995 in type I diabetes mellitus

The effect of a new long-acting somatostatin analog SMS 201-995 (SMS) on hormonal mechanisms controlling the glucose metabolism was tested in 8 type I diabetics over a 3-day period. In addition to dietary measures and conventional insulin therapy, the patients received a subcutaneous dose of 50 micrograms SMS three times daily for 3 days. Serum growth hormone (GH) was measured at various intervals throughout the investigational period. Glucagon, somatomedin C (SM-C), triiodothyronine, thyroxine, luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin (PRL) were also determined before and at the end of the therapy with SMS. Basal GH and plasma SM-C had decreased significantly (p less than 0.05 and p less than 0.01, respectively) by the 3rd day. In all cases the insulin requirements could be reduced (mean 28%) without deterioration of the metabolic control. Moreover, blood glucose profiles showed a tendency to lower postprandial peaks after SMS treatment. Glucagon, triiodothyronine, thyroxine, LH, FSH and PRL showed no significant changes. No side effects or alterations in laboratory chemistries were recorded. Dampening of glucose oscillations and counterregulatory mechanisms, and reduction of insulin dosage by SMS may enable a better control of unstable diabetes. Its slow plasma clearance and long action compared to the native peptide will warrant the use of this analog as a additive to standard diabetes therapy in more prolonged trials.     

Treatment of inappropriate secretion of thyrotropin with somatostatin analog SMS 201-995

Inappropriate thyrotropin secretion (IST) may originate from either neoplastic disease (nIST) or non-neoplastic resistance to thyroid hormone (nnIST). An inhibitory effect of somatostatin on TSH secretion has been documented. In an attempt to elucidate the possible therapeutic effect of this peptide on nIST and nnIST, a study was conducted in 7 such patients. Sandostatin (SMS 201-995) was administered in daily doses of 100 micrograms for several days to 1 month. Four patients with nIST responded with a fall in circulating TSH as well as alpha-subunit with concomitant normalization of free thyroxine and clear symptomatic improvement. In the 3 nnIST patients this effect was considerably less apparent and a partial TSH escape was observed on long-term treatment in 2 cases. The importance of somatostatin and its analogs in the management of thyroid malignancy is stressed.     

Unexpected growth-stimulatory effect of somatostatin analogue on cultured human pancreatic carcinoid cells.

Therapeutic efficacy of a synthetic somatostatin analogue for the treatment of carcinoid tumors is still controversial. In vivo studies performed in our laboratory showed that a somatostatin analogue, SMS 201-995, significantly inhibited growth of human pancreatic carcinoid (BON) tumors xenotransplanted into athymic nude mice. In the present study, however, SMS 201-995 did not inhibit in vitro growth of BON cells, but rather SMS 201-995 stimulated growth in a dose-dependent fashion. The growth-stimulatory effect was likely mediated through the reduction of cyclic AMP production. Unsuccessful treatment of certain types of carcinoid tumor with SMS 201-995 may be partly due to the direct growth-stimulatory effect of SMS 201-995 on carcinoid cells.     

Somatostatin analog treatment of acromegaly: new aspects

Ten acromegalics received daily doses of 200-300 micrograms of a long-acting somatostatin analog, SMS 201-995 (Sandostatin, SMS), for an average of 64 weeks. Basal mean GH values of 44 +/- (SE) 7.8 ng/ml had fallen into the normal range at the end of the observation period (mean 64 weeks). This effect was accompanied by a substantial drop in somatomedin-C values. Reduction of pituitary tumor size could be documented in 3 of 6 patients. Whereas SMS did not affect high plasma PRL in 4 microprolactinoma patients, lactotrophs turned sensitive to this agent in mixed GH/PRL tumors. In a comparative study between SMS and bromocriptine, the former normalized circulating GH in 10 of 17 acromegalics in an acute trial, whereas bromocriptine was effective in only 5. A combination of both substances was effective in 2 of 3 patients who were insensitive to single drug administration. Cultures of GH-secreting tumor cells showed a statistically significant hormone decrease in the medium when exposed to SMS. However, in some instances, a diminution of the GH contents of the tumor cells was also observed, presumably as the basis for intracellular breakdown and clinical tumor shrinkage.     

Plasma alpha-subunit levels during the treatment of pituitary adenomas with the somatostatin analog (SMS 201-995)

The effect os SMS 201-995 (Sandostatin), a long-acting somatostatin analog, on different types of pituitary adenomas including alpha-subunit elevation is illustrated in this report. Treatment induced a fall in hCG levels in a woman with a pituitary adenoma producing only alpha-subunit. In 3 acromegalic patients, there was only a partial drop in GH and alpha-hCG. The same effect was observed in a woman with menopausal FSH and LH levels. SMS reduced plasma TSH and alpha-hCG in a case of thyrotropic adenoma. Two patients exhibiting FSH- and alpha-hCG-secreting adenomas did not respond to acute administration of SMS 201-995. More patients have to be treated before a definitive statement can be made on the usefulness of somatostatin analogs in the management of different types of pituitary adenomas.     

Effect of sandostatin on CRF-stimulated secretion of ACTH, beta-lipotropin and beta-endorphin.

The influence exerted by somatostatin on the secretion of ACTH and opioid peptides has still to be clarified. To gain further information on this issue, we performed in 10 normal volunteers two CRF tests (100 micrograms i.v.) one of which was preceded by s.c. injection of 100 micrograms of the long-acting somatostatin analogue SMS 201-995 (Sandostatin, Sandoz) (SMS), given 30 minutes before CRF. Premedication with SMS markedly inhibited the response of beta-EP to CRF, leaving unchanged the response of beta-LPH, ACTH and cortisol; mean incremental areas of beta-EP were 199.8 +/- 49.31 (SEM) vs 532.9 +/- 95.91 pmol 120 min (P less than 0.01) in the CRF test with and without SMS, respectively. To interpret the selective inhibitory effect of SMS on CRF-stimulated beta-EP secretion, it can be hypothesized that: a) the action of SMS was confined to a population of pituicytes preferentially secreting beta-EP; b) SMS interfered with the processing of POMC inhibiting the formation of beta-EP; c) SMS acted on extrapituitary, possibly peripheral, sources of beta-EP. In conclusion, this study indicates that, in man, somatostatin selectively inhibits the CRF-induced secretion of beta-EP, but not that of ACTH and beta-LPH, by an action that may be exerted at pituitary or extrapituitary level. This is a further example of dissociated secretion of POMC-derived peptides.     

Somatostatin structure-activity studies in the stomach

Somatostatin may inhibit gastric exocrine functions independent of blockade of gastrin secretion. In order to further investigate this suppressive effect, somatostatin derivatives were injected to cats bearing a cannulated gastric fistula under pentagastrin stimulation. Results showed that somatostatin-14 was more potent than somatostatin-28 in this particular model. Analogues with substituted residues exhibited a variable spectrum of actions on hormone release and gastric function. A cyclic pentapeptide was deprived of gastric or GH inhibitory properties whereas the related peptide with a benzyl-protecting group on Thr was only devoid of gastric effect. The octapeptide SMS 201-995 was described as a potent inhibitor of gastric secretion in comparison with natural somatostatin in rats and also in humans, but was unable to induce maximal suppression of acid output in the cat model. Differences in gastric effect of different derivatives could be explained on the basis of binding to a selective subset of receptors, since at least two binding sites have been identified in the stomach mucosa. Serial studies with short cyclic somatostatin should help to establish a clear relationship between peptide structure and inhibition of gastric secretion.     

Two amino acids, located in transmembrane domains VI and VII, determine the selectivity of the peptide agonist SMS 201-995 for the SSTR2 somatostatin receptor.

Human somatostatin receptor subtypes (SSTR1-5) bind their natural ligands SRIF-14 and SRIF-28 with high affinity. By contrast, short synthetic SRIF analogues such as SMS 201-995, a peptide agonist used for the treatment of various endocrine and malignant disorders, display sub-nanomolar affinity only for the receptor subtype SSTR2. To understand the molecular nature of selective peptide agonist binding to somatostatin receptors we have now, by site-directed mutagenesis, identified amino acids mediating SMS 201-995 specificity for SSTR2. Sequentially, amino acids in SSTR1, a receptor subtype exhibiting low affinity for SMS 201-995, were exchanged for the corresponding SSTR2 residues. After three consecutive steps, in which eight amino acids were exchanged, a SSTR1 mutant receptor with high affinity for SMS 201-995 was obtained. Receptor mutants with different combinations of these eight amino acids were then constructed. A single Ser305 to Phe mutation in TM VII increased the affinity of SSTR1 for SMS 201-995 nearly 100-fold. When this mutation was combined with an exchange of Gln291 to Asn in TM VI, almost full susceptibility to SMS 201-995 was obtained. Thus, it is concluded that the specificity of SMS 201-995 for SSTR2 is mainly defined by these two amino acids in transmembrane domains VI and VII. Using the conjugate gradient method we have, by analogy to the well established structure of bacteriorhodopsin, built a model for SRIF receptor-ligand interactions that explains the importance of Gln291 and Ser305 for the selectivity of agonists.     

Somatostatin analog (SMS 201-995) inhibits the basal and angiotensin II-stimulated 3H-thymidine uptake by rat adrenal glands

The effects of a long-acting somatostatin analog SMS 201-995 injections on the basal and angiotensin II-stimulated [3H]-thymidine uptake by the rat adrenal glands incubated in vitro were examined. It was shown that SMS 201-995 significantly inhibited the [3H]-thymidine uptake and, additionally, suppressed the stimulatory effect of a single angiotensin II injection.     

Improvement of diabetes after treatment with somatostatin analogue SMS 201-995 in an acromegalic patient

The beneficial effects of long acting somatostatin analogue SMS 201-995 in an acromegalic patient affected by severe diabetes mellitus are reported. Neither human insulin alone nor human insulin plus bromocriptine allowed satisfactory metabolic control though, with the latter treatment, virtually normal plasma GH levels were reached. Conversely, addition of SMS 201-995 to insulin treatment led to normalization of blood glucose. This result was obtained with a dose of SMS 201-995 of 400 micrograms/day and only after 3 weeks of therapy.     

Interactions between neuropeptides and dexamethasone on the mitogenic response of rabbit spleen lymphocytes.

The interactions between vasoactive intestinal peptide (VIP), substance P (SP), a somatostatin analog (SMS 201-995) and dexamethasone have been investigated on the Con A mitogenic response of rabbit spleen cells. The neuropeptide regulatory effects appeared to be time dependent: when added with the Con A mitogen, they inhibited (VIP) or did not modulate (SMS and SP) the rabbit lymphocyte proliferation and did not change the inhibitory effect induced by a dexamethasone preincubation. When added 18 h before the mitogen, they all induced an increase of the proliferative response at high concentration. The mitogenic response observed when adding dexamethasone to lymphocytes previously preincubated in the presence of neuropeptides was not different from control response except with SMS 10(-10) M. The similar lymphocyte responses obtained whatever the neuropeptide suggested that the immunomodulatory effect induced by a neuropeptide preincubation might be mediated by the induction of common effector(s).     

SMS 201-995: a very potent and selective octapeptide analogue of somatostatin with prolonged action

Stepwise modification of a conformationally stabilised analogue of the fragment of somatostatin which had been thought to be essential biologically active moiety has enabled us to synthesise the analogue H-(D) Phe-Cys-Phe-(D) Trp-Lys-Thr-Cys-Thr(ol) code-named SMS 201-995, which in vitro is three times more potent than the native hormone in inhibiting the secretion of growth hormone, which is highly resistant to degradation by pure enzymes and by tissue homogenates, which in vivo in rat and rhesus monkey is (depending on test system) at least 20 times more active than somatostatin, which is much longer acting, and which moreover in both species is much more selective in inhibiting the secretion of growth hormone than that of insulin. The compound is active by several routes of administration including the oral, is well tolerated both in laboratory animals and in man, and is currently undergoing preliminary clinical trial.     

Effects of a long-acting somatostatin analogue on pituitary-adrenocortical secretion in normal human subjects

A potent and long-acting somatostatin analogue, SMS 201-995 (SMS) is currently employed for the treatment of various diseases with hypersecretion of hormones such as acromegaly and gastrinoma. The suppressive effects of SMS are also reported on the other pituitary and gastrointestinal hormones. The corticotropic-adrenocortical axis is a crucial hormonal complex in maintaining normal activity and life itself. In this study, the effects of SMS on corticotropic-adrenocortical functions were studied, since the effects of SMS on this hormonal axis are not well established. Seven normal males received a sc injection of 100 micrograms SMS or placebo at 0830 h and 100 micrograms synthetic human corticotropin-releasing hormone (hCRH) intravenously (SMS-hCRH study). Five of the 7 subjects were given an injection of a synthetic (1-24) ACTH (250 micrograms or 63 micrograms) at 0900 h after 100 micrograms SMS or a placebo at 0830 h (SMS-ACTH study). Blood samples were drawn at -30, 0, 15, 30, 60, 90 and 120 min after the hCRH injection for the determination of ACTH and cortisol in the SMS-hCRH study, and cortisol and aldosterone in the SMS-ACTH study. Although significant rises in plasma ACTH and cortisol levels were observed regardless of the preinjection of SMS, their responses to hCRH were significantly lower with the pretreatment with SMS than without SMS. A significant increase in plasma cortisol and aldosterone was observed in response to synthetic ACTH with both ACTH alone and the combined administration of SMS and ACTH, at either dose of ACTH. However, no significant difference in cortisol and aldosterone secretion was detected with and without SMS.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of subcutaneous injection of a long-acting analogue of somatostatin (SMS 201-995) on plasma thyroid-stimulating hormone in normal human subjects

SMS 201-995 (SMS), a synthetic analogue of somatostatin (SRIF) has been shown to be effective in the treatment of the hypersecretion of hormones such as in acromegaly. However, little is known about the effects of SMS on the secretion of thyroid-stimulating hormone (TSH) in normal subjects. In this study, plasma TSH was determined with a highly sensitive immunoradiometric assay, in addition to the concentration of SMS in plasma and urine with a radioimmunoassay, following subcutaneous injection of 25, 50, 100 micrograms of SMS (4 subjects/dose) or a placebo (6 subjects) to normal male subjects, at 0900 h after an overnight fast. The plasma concentrations of SMS were dose-responsive and the peak levels were 1.61 +/- 0.09, 4.91 +/- 0.30 and 8.52 +/- 1.18 ng/ml, which were observed at 30, 15 and 45 min after the injection of 25, 50 and 100 micrograms of SMS, respectively. Mean plasma disappearance half-time of SMS was estimated to be 110 +/- 3 min. Plasma TSH was suppressed in a dose dependent manner and the suppression lasted for at least 8 hours. At 8 hours after the injection of 25, 50 and 100 micrograms of SMS, the plasma TSH levels were 43.8 +/- 19.4, 33.9 +/- 9.4 and 24.9 +/- 3.2%, respectively, of the basal values. The results suggest that SMS suppresses secretion of TSH from the normal thyrotrophs in man and thus also that attention should be paid to possible hypothyroidism during the long-term treatment of patients such as those with acromegaly with this potent analogue of SRIF.     

Insulin-induced hypoglycemia, L-dopa and arginine stimulate GH secretion through different mechanisms in man

We sought to clarify the mechanisms of growth hormone (GH) secretion induced by insulin hypoglycemia, L-dopa, and arginine in man. The secretion of GH as measured by increased plasma level, in response to oral administration of 500 mg L-dopa or 30 min-infusion of arginine, was not modified by prior intravenous administration of 200 micrograms GH-releasing hormone (GHRH). It was, however, completely blocked by preadministered 50 micrograms SMS201-995, a long-acting somatostatin (SRIH) analog. GH release with 200 micrograms GHRH was completely blocked by 100 micrograms SMS201-995. GH secretion caused by insulin-induced hypoglycemia was significantly reduced but still present after administration of 100 micrograms of the analog. These results suggest that a suppression of SRIH release may be partially involved in the stimulatory mechanism of GH secretion by L-dopa. Coadministration of GHRH accentuated the stimulatory effect of arginine on GH secretion. Arginine significantly raised plasma TSH levels. These findings suggest that arginine suppresses SRIH release from the hypothalamus to cause GH secretion because SRIH suppresses TSH secretion. It is also suggested that some factor (or factors) other than GHRH and SRIH are involved in the mechanism by which insulin-induced hypoglycemia stimulates GH secretion, because the effect of insulin was not fully blocked in the presence of SRIH analog. Thus all the tests for GH release appear to act via different mechanisms.     

111In-DTPA-D-Phe1]-octreotide, a potential radiopharmaceutical for imaging of somatostatin receptor-positive tumors: synthesis, radiolabeling and in vitro validation.

Somatostatin receptor-positive human tumors can be detected using radioiodinated analogues of somatostatin, both in vitro and in vivo. [123I-Tyr3]-octreotide has been successfully used in the visualization of somatostatin receptor-positive tumors by gamma camera scintigraphy, but this radiopharmaceutical has some major drawbacks, which can be overcome with other radionuclides such as 111In. As starting material for a potentially convenient radiopharmaceutical, a diethylenetriaminopentaacetic acid (DTPA) conjugated derivative of octreotide (SMS 201-995) was prepared. This peptide, [DTPA-D-Phe1]-octreotide (SDZ 215-811) binds more than 95% of added 111In in an easy, single-step labeling procedure without necessity of further purification. The specific somatostatin-like biologic effect of these analogues was proven by the inhibition of growth hormone secretion by cultured rat pituitary cells in a dose-dependent fashion by octreotide, [DTPA-D-Phe1]-octreotide and non-radioactive [115In-DTPA-D-Phe1]-octreotide. The binding of [111In-DTPA-D-Phe1]-octreotide to rat brain cortex membranes proved to be displaced similarly by natural somatostatin as well as by octreotide, suggesting specific binding of [111In-DTPA-D-Phe1]-octreotide to somatostatin receptors. The binding of the indium-labeled compound showed a somewhat lower affinity when compared with the iodinated [Tyr3]-octreotide, but indium-labeled [DTPA-D-Phe1]-octreotide still binds with nanomolar affinity. In conjunction with in vivo studies, these results suggest that [111In-DTPA-D-Phe1]-octreotide is a promising radiopharmaceutical for scintigraphic imaging of somatostatin receptor-positive tumors.     

Delta 6- and delta 12-desaturase activities and phosphatidic acid formation in microsomal preparations from the developing cotyledons of common borage (Borago officinalis).

The somatostatin receptors on rat pancreatic acinar membranes were demonstrated by use of a radioiodinated (125I-) analogue of somatostatin (SMS 204-090 or [Tyr3]SMS). The tracer was found to bind to the receptor with a Kd of 58 pM. The number of sites detected by this tracer (4.7 pmol/mg of protein) was 5-10 times higher than the number of sites previously found with other tracers. Since the level of non-specific binding was also very low as compared with findings with other tracers, 125I-204-090 might be of interest in future attempts to characterize the somatostatin receptors in the pancreas. The prelabelled membranes were solubilized with 1% CHAPS, and the solubilized complexes were found to adsorb to wheat-germ-agglutinin-coupled agarose, from which they could be eluted with 4 mM-triacetylchitotriose. The complexes within this eluate were shown by gel filtration on Trisacryl GF-2000 to have an Mr of about 400,000. The dissociation of the complexes was augmented both within the membranes as well as in the solubilized state by incubation with the GTP analogue guanosine 5'-[gamma-thio]triphosphate, indicating that the complexes are probably functionally linked to a guanine-nucleotide-binding regulatory protein. After SDS/slab-gel electrophoresis and autoradiography of cross-linked complexes after treatment with the heterobifunctional reagent N-5-azido-2-nitrobenzoyloxysuccinimide, a broad band occurred at approximately Mr 90,000 both in the membranes and in the eluates of complexes after lectin-adsorption chromatography. We conclude that the augmentation of the number of detectable sites for binding of somatostatin, as well as the very low level of non-specific binding obtained by the use of 125I-[Tyr3]SMS as tracer, has made it possible for us to demonstrate the solubilization of the somatostatin receptor in conjunction with its ligand and a GTP-binding regulatory protein, and we have succeeded in cross-linking 125I-[Tyr3]SMS to a binding subunit of Mr 90,000 in the membranes and in demonstrating the presence of the same labelled binding subunit within complexes solubilized and chromatographed on a lectin column before cross-linking.     

Backbone conformations and side chain flexibility of two somatostatin mimics investigated by molecular dynamics simulations

Molecular dynamics simulations with two designed somatostatin mimics, SOM230 and SMS 201‐995, were performed in explicit water for a total aggregated time of 208 ns. Analysis of the runs with SOM230 revealed the presence of two clusters of conformations. Strikingly, the two sampled conformers correspond to the two main X‐ray structures in the asymmetric unit of SMS 201‐995. Structural comparison between the residues of SOM230 and SMS 201‐995 provides an explanation for the high binding affinity of SOM230 to four of five somatostatin receptors. Similarly, cluster analysis of the simulations with SMS 201‐995 shows that the backbone of the peptide interconverts between its two main crystallographic conformers. The conformations of SMS 201‐995 sampled in the two clusters violated two different sets of NOE distance constraints in agreement with a previous NMR study. Differences in side chain fluctuations between SOM230 and SMS 201‐995 observed in the simulations may contribute to the relatively higher binding affinity of SOM230 to most somatostatin receptors. Proteins 2009. © 2008 Wiley‐Liss, Inc.