| 1331448 |
Development of a novel class of cyclic hexapeptide oxytocin antagonists based on a natural product |
10.1021/jm00099a019. |
J Med Chem |
Development of a novel class of cyclic hexapeptide oxytocin antagonists based on a natural product
Abstract
- A new structural class of cyclic hexapeptide oxytocin antagonists derived from Streptomyces silvensis and typified by L-365,209 (cyclo-L-prolyl1-D-phenylalanyl2-L- isoleucyl3-D-dehydropiperazyl4-L-dehydroperazyl5-D-(N- methyl)phenylalanyl6) was recently reported. In this paper we further delineate the structure-activity profile for this new class by systematic study of L-365,209 analogs obtained by total synthesis. The optimal combination of cyclic amino acid ring sizes at positions 1, 4, and 5 and the role of the N-alkyl substituent at position 6 was elucidated. The lipophilic amino acids at positions 2 and 3 and the unusual amino acid D-dehydropiperazic acid at position 4 were found to be the most critical residues for obtaining good oxytocin receptor affinity. Analogs containing a basic side chain at the less critical 5- and 6-positions maintained good receptor affinity and also had useful levels of water solubility for intravenous formulation. By combining potency- and solubility-enhancing substitutions, several analogs were identified that have the desired combination of properties in vitro (22, cyclo-L-prolyl-D-tryptophanyl-L-isoleucyl-D-pipecolyl-L-pipeco lyl-D- histidyl; 25, cyclo-L-prolyl-D-2-naphthylalanyl-L-isoleucyl-D-pipecolyl-L -pipecolyl-D- histidyl; 26, cyclo-L-prolyl-D-tryptophanyl-L-isoleucyl-D-dehydropiperazyl-L-++ pipecolyl-D-histidyl; 33, cyclo-L-prolyl-D-tryptophanyl-L-isoleucyl-D-pipecolyl-L- piperazinylcarboxy-D-(N-methyl)phenylalanyl; 34, cyclo-L-prolyl-D-phenylalanyl-L-isoleucyl-D-dehydropiperazyl-L-or nithyl- D-(N-methyl)phenylalanyl). In general, this class exhibited good selectivity for binding to the oxytocin receptor versus the arginine vasopressin V1a and V2 receptor subtypes, although increased V2 receptor affinity was observed in one case (32, cycloL-prolyl-D-2-naphthylalanyl-L-isoleucyl-D-pipecolyl-L- lysyl-D-(N- methyl)phenylalanyl). Unexpectedly, compound 33 was found to stimulate contractions of the isolated rat uterus via activation of the uterine bradykinin receptor. Compounds 22, 25, 26, 33, and 34 were found to be potent antagonists of oxytocin-stimulated contraction of the rat uterus in vitro and in vivo. Compounds 22 and 25 were additionally characterized as potent antagonists of oxytocin-stimulated uterine contractions in the near-term pregnant rhesus monkey. These studies thus demonstrate the selectivity and efficacy of certain members of this novel class of antagonists and suggest their use as pharmacological tools in further defining the role of oxytocin in both term and preterm labor.
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| 1332703 |
Detection of topoisomerase I gene point mutation in CPT-11 resistant lung cancer cell line |
10.1016/0006-291x(92)91094-7. |
Biochem Biophys Res Commun |
Detection of topoisomerase I gene point mutation in CPT-11 resistant lung cancer cell line
Abstract
- CPT-11, a recently developed topoisomerase I (Topo I) inhibitor, attracts the attention not only of basic researchers but also of clinicians because of its high antitumor activity. The CPT-11 resistant human lung cancer cell line, PC-7/CPT, showed 10-fold resistance compared to parental cell line, PC-7. The total activity of Topo I in the resistant cell line was one fourth that of the parental sensitive cell line. The Topo I from the resistant cells was also 5-fold more resistant to the inhibitory effect of CPT-11 than that of the parental cells. We speculated that the alteration of the Topo I gene may be responsible for the change in topoisomerase activity of the CPT-11 resistant cell line. Therefore, we analyzed the mutation of Topo I gene using the method of single strand conformation polymorphism of polymerase chain reaction and the reverse transcriptase. We divided Topo I cDNA into ten fragments which overlapped each other and covered whole coding sequences of the Topo I cDNA. We observed mobility shift of two fragments in the PC-7/CPT, suggesting the presence of some mutations in these fragments. We performed the direct-sequencing of these portions by the dideoxy chain termination method and observed an altered sequence having a G to A base change in PC-7/CPT. This base substitution results in replacement of the conserved threonine at 729 position with alanine. These results suggest that the point mutation of Topo I gene is related to the decreases of Topo I activity and the sensitivity to Topo I inhibitor in PC-7/CPT cells.
|
| 1337145 |
Somatostatin receptors, an expanding gene family: cloning and functional characterization of human SSTR3, a protein coupled to adenylyl cyclase. |
10.1210/mend.6.12.1337145 |
Mol. Endocrinol. |
Somatostatin receptors, an expanding gene family: cloning and functional characterization of human SSTR3, a protein coupled to adenylyl cyclase.
Abstract
- We previously reported the cloning of two distinct somatostatin receptor (SSTR) subtypes, SSTR1 and SSTR2. Although both SSTR1 and SSTR2 bound somatostatin specifically and with high affinity, neither was coupled to adenylyl cyclase, a major cellular effector of somatostatin's actions. Here we report the cloning and functional characterization of a third member of the SSTR family. Human SSTR3 is a protein of 418 amino acids and has 45% and 46% identity with human SSTR1 and SSTR2, respectively. RNA blotting studies showed that SSTR3 mRNA could be readily detected in brain and pancreatic islets. The pharmacological properties of human SSTR3 were characterized by transiently expressing the human SSTR3 gene in COS-1 cells. Membranes from cells expressing human SSTR3 bound the somatostatin agonist [125I]CGP 23996 specifically and with high affinity, with a rank order of potency of somatostatin-28 = CGP 23996 > somatostatin-14 > SMS-201-995. Studies using cells transiently coexpressing the human dopamine D1 receptor and human SSTR3 showed that somatostatin was able to inhibit dopamine-stimulated cAMP formation in a dose-dependent manner, indicating that SSTR3 was functionally coupled to adenylyl cyclase. These
Results indicate that the diverse biological effects of somatostatin are mediated by a family of receptor with distinct, but overlapping, tissue distributions, unique pharmacological properties, and potentially different functions.
|
| 1337381 |
Characterization of the novel CCK analogs JMV-180, JMV-320, and JMV-332 in H345 cells |
10.1016/0196-9781(92)90033-y. |
Peptides |
Characterization of the novel CCK analogs JMV-180, JMV-320, and JMV-332 in H345 cells
Abstract
- The interaction of the novel CCK analogs JMV-180, JMV-320, and JMV-332 with CCK-B/gastrin receptors on small cell lung cancer (SCLC) cells was investigated. JMV-180, JMV-320, and JMV-332 potently inhibited specific binding of 125I-CCK-8 to CCK-B/gastrin receptors expressed on the SCLC cell line NCI-H345 (H345) with IC50 values of 4.9, 1.8, and 7.0 nM, respectively. JMV-320 and JMV-332 stimulated intracellular calcium (Ca2+i) release in a dose-dependent manner in cells preloaded with indo-1. JMV-180 did not stimulate Ca2+i but inhibited the Ca2+i release elicited by 10 nM CCK-8 in a dose-dependent manner. These data indicate that JMV-320 and JMV-332 function as CCK-B/gastrin receptor agonists while JMV-180 functions as a CCK-B/gastrin receptor antagonist in H345 cells.
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| 1340179 |
Immunosuppressive activity of threonine-containing analogues of cyclolinopeptide A |
None |
Arch Immunol Ther Exp (Warsz) |
Immunosuppressive activity of threonine-containing analogues of cyclolinopeptide A
Abstract
|
| 1344901 |
Release of heptapeptide toxin (microcystin) during the decomposition process of Microcystis aeruginosa |
10.1002/nt.2620010110. |
Nat Toxins |
Release of heptapeptide toxin (microcystin) during the decomposition process of Microcystis aeruginosa
Abstract
- The decomposition process of toxic blue-green alga (cyanobacteria), Microcystis aeruginosa, under dark and aerobic condition was investigated in relation to the change of the amounts of heptapeptide toxins (microcystins YR and LR) by two experiments: one with Microcystis cells and the other with two purified microcystins. In the experiment with Microcystis cells, an increase of heterotrophic bacteria observed from the beginning of the experiment, was followed by decomposition of the algal cells and the subsequent release of microcystins into the filtrate fraction. The amounts of the toxins initially present in the cells were quantitatively detected in the filtrate fraction on the 35th day. The decomposition of microcystin YR began on the 42nd day. The decomposition rate of the two toxins was different. The decomposition rate of purified microcystins YR and LR, compared in distilled water and culture medium, respectively, indicated clearly that microcystin YR was more labile to decomposition than microcystin LR in the culture medium. At the end of the experiment (45th day) microcystin YR decreased to 58.6%, while 86.2% of microcystin LR remained.
|
| 1346068 |
Cloning and functional characterization of a family of human and mouse somatostatin receptors expressed in brain, gastrointestinal tract, and kidney. |
10.1073/pnas.89.1.251 |
Proc. Natl. Acad. Sci. U.S.A. |
Cloning and functional characterization of a family of human and mouse somatostatin receptors expressed in brain, gastrointestinal tract, and kidney.
Abstract
- Somatostatin is a tetradecapeptide that is widely distributed in the body. It acts on multiple organs including brain, pituitary, gut, exocrine and endocrine pancreas, adrenals, thyroid, and kidneys to inhibit release of many hormones and other secretory proteins. In addition, it functions as a neuropeptide affecting the electrical activity of neurons. Somatostatin exerts its biological effects by binding to specific high-affinity receptors, which appear in many cases to be coupled to GTP-binding proteins. Here we report the cloning, functional expression, and tissue distribution of two different somatostatin receptors (SSTRs). SSTR1 and SSTR2 contain 391 and 369 amino acids, respectively, and are members of the superfamily of receptors having seven transmembrane segments. There is 46% identity and 70% similarity between the amino acid sequences of SSTR1 and SSTR2. Stably transfected Chinese hamster ovary cells expressing SSTR1 or SSTR2 exhibit specific somatostatin binding, with an apparently higher affinity for somatostatin-14 than somatostatin-28, and NH2-terminally extended form of somatostatin-14. RNA blotting studies show that SSTR1 and SSTR2 are expressed at highest levels in jejunum and stomach and in cerebrum and kidney, respectively. A SSTR1 probe hybridized to multiple DNA fragments in EcoRI digests of human and mouse DNA, indicating that SSTR1 and SSTR2 are members of a larger family of somatostatin receptors. Thus, the biological effects of somatostatin are mediated by a family of receptors that are expressed in a tissue-specific manner.
|
| 1346648 |
Differential coupling of somatostatin1 receptors to adenylyl cyclase in the rat striatum vs. the pituitary and other regions of the rat brain |
None |
J Pharmacol Exp Ther |
Differential coupling of somatostatin1 receptors to adenylyl cyclase in the rat striatum vs. the pituitary and other regions of the rat brain
Abstract
- Subtypes of somatostatin (SRIF) receptors are expressed in the rat brain and may mediate the diverse actions of SRIF. In the present study we show that subtypes of SRIF receptors in different regions of the rat brain are differentially sensitive to the cyclic hexapeptide SRIF analog, MK 678. SRIF1 receptors are sensitive to MK 678 and found in high density in the cortex, hippocampus and striatum, as well as in the anterior pituitary. The pituitary appears to express only the SRIF1 receptor. The cortex, hippocampus and striatum also express SRIF2, or MK 678-insensitive, receptors. The proportion of SRIF1 receptors varies in different brain regions. In the cortex and hippocampus, SRIF1 receptors comprise approximately 50% of the total SRIF receptor population, whereas in the striatum SRIF1 receptors comprise the majority (86%) of SRIF receptors. SRIF1 receptors in the pituitary, cortex and hippocampus mediate, at least in part, the ability of SRIF to inhibit forskolin-stimulated adenylyl cyclase activity as MK 678 produced significant inhibition of activity in these tissues. However, in the striatum, MK 678 had no significant effect on forskolin-stimulated adenylyl cyclase activity, despite a significant inhibition produced by SRIF. The specific labeling of these receptors in the striatum by 125IMK 678 is abolished in the presence of high concentrations of the nonhydrolyzable GTP analog, GTP gamma S, suggesting that SRIF1 receptors in this brain region are coupled to G proteins. The SRIF1 receptors in the striatum may be coupled via G proteins to cellular transducing systems other than adenylyl cyclase.
|
| 1346715 |
Characterization of 125ITyr11-somatostatin binding sites in the rabbit retina |
10.1016/0143-4179(92)90148-p. |
Neuropeptides |
Characterization of 125ITyr11-somatostatin binding sites in the rabbit retina
Abstract
- We have identified specific receptors for somatostatin (SS) in the rabbit retina using the radioligand 125ITyr11-Somatostatin. 125ITyr11-SS bound with high affinity to retinal membranes as was ascertained by both kinetic and saturation experiments. Scatchard analysis of the saturation data for 125ITyr11-SS binding to retinal membranes suggest a single population of sites with an apparent affinity constant (KD) of 0.90 +/- 0.20 nM and a maximum number of binding sites (Bmax) of 104 +/- 52 fmol/mg protein. The specific binding of 125ITyr11-SS was displaced in a dose-dependent manner by SS, Tyr11-SS, SMS 201-995, SS-28 and D-Trp8-SS. The inactive SS analog SS28(1-14) as well as the peptides CRF and bombesin had no effect. In addition, the specific binding of 125ITyr11-SS was attenuated by GTPgS. These findings demonstrate the presence of a selective receptor for SS in the rabbit retina that is coupled to guanine nucleotide binding protein(s).
|
| 1349838 |
Detection of a single base substitution of the gene for prothrombin Tokushima. The application of PCR-SSCP for the genetic and molecular analysis of dysprothrombinemia |
None |
Int J Hematol |
Detection of a single base substitution of the gene for prothrombin Tokushima. The application of PCR-SSCP for the genetic and molecular analysis of dysprothrombinemia
Abstract
- The genetic and molecular basis of a mutant prothrombin of 'prothrombin Tokushima' was studied by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and PCR-restriction fragment length polymorphism (PCR-RFLP) analyses. The abnormal gene was detected by altered migration by PCR-SSCP and by the loss of an MspI site by PCR-RFLP. The gene for prothrombin Tokushima was shown to be inherited from the mother of the proband. Sequencing analysis using PCR-amplified genomic DNA clarified a substitution of thymine (T) for cytosine (C) at position 9,490, changing arginine (Arg) to tryptophan (Trp) at position 418 of the polypeptide chain. This point mutation is assumed to be the molecular basis of prothrombin Tokushima, firstly, because of the absence of distinct changes in Southern blot analysis of the proband's DNA (using a full-length human prothrombin cDNA as a probe), secondly, because it has the same molecular weight as the abnormal gene product, and, thirdly, because of the absence of other amino acid abnormalities in the proteolytic peptide-fragments. It is concluded that PCR-SSCP and PCR-RFLP were useful for detecting the abnormal gene and for directly diagnosing the carrier status of dysprothrombinemia. This is the first report of gene analysis of dysprothrombinemia.
|
| 1351397 |
Light microscopic radioautographic localization of somatostatin binding sites in the brainstem of the rat |
10.1016/0891-0618(92)90035-o. |
J Chem Neuroanat |
Light microscopic radioautographic localization of somatostatin binding sites in the brainstem of the rat
Abstract
- The distribution of somatostatin binding sites was investigated by light microscopic radioautography in the brainstem of the rat following in vitro labeling with 125I-Tyr0-DTrp8-somatostatin14. Moderate to high labeling densities were detected within the superior colliculus, the locus coeruleus and subcoeruleus, the parabrachial complex, the nucleus of the solitary tract and the dorsal motor nucleus of the vagus. Most of the white matter was devoid of specific somatostatin binding except for fibers of the glossopharyngeal nerve and the spinal trigeminal tract. In most of the labeled areas, 125I-somatostatin binding was evenly distributed between neuropil and perikarya. In a few instances, however, the binding clearly predominated over nerve cell bodies: namely in the dorsal motor nucleus of the vagus and in the pontine and medullary tegmentum. In the latter two regions, the labeled neurons were identified in adjacent sections by tyrosine hydroxylase immunohistochemistry as belonging to the A5 and A1 catecholamine cell groups, respectively. These findings, together with the confirmed association of somatostatin binding sites with noradrenergic neurons in the locus coeruleus, suggest that interactions with catecholaminergic systems may represent a major mode of action for somatostatin in the brainstem.
|
| 1352176 |
Somatostatin receptor elevation in rat striatum after diisopropylfluorophosphate administration |
10.1016/0361-9230(92)90097-h. |
Brain Res Bull |
Somatostatin receptor elevation in rat striatum after diisopropylfluorophosphate administration
Abstract
- The acute and chronic administration of diisopropylfluorophosphate (DFP), an inhibitor of acetylcholinesterase or of atropine, a blocker of muscarinic cholinergic receptors, did not affect somatostatin-like immunoreactivity (SLI) content in the striatum of rats. Acute and chronic DFP administration increased the number of specific 125I-Tyr11-somatostatin (125I-Tyr11-SS) receptors in cells dissociated from the striatum without changing the affinity constant. Although the increase could be blocked by pretreatment with atropine, it was not due to a direct effect by DFP on somatostatin (SS) receptors, because no rise in 125I-Tyr11-SS binding was produced by high concentrations of DFP (10(-5) M) when added in vitro. The acute administration of atropine alone had no observable effect on the number of SS receptors. However, repeated atropine administration produced a significant decrease in the 125I-Tyr11-SS binding in cells dissociated from the striatum, although the affinity constant was unchanged. The results suggest that interactions between somatostatinergic and cholinergic receptors may be of importance in the rat striatum.
|
| 1352707 |
Evidence that endogenous somatostatin (SRIF) exerts a tonic inhibitory effect on the rat renin--angiotensin--aldosterone system |
None |
In Vivo |
Evidence that endogenous somatostatin (SRIF) exerts a tonic inhibitory effect on the rat renin--angiotensin--aldosterone system
Abstract
- The bolus ip. administration of a SRIF antagonist (SRIF-A) (60 nM/rat) significantly increased renin activity (PRA) and plasma aldosterone concentration (PAC) in rats, without affecting natremia, kalaemia and the blood levels of ACTH or corticosterone. SRIF-A also raised PAC in rats whose renin-angiotensin system had been pharmacologically interrupted by combined captopril/angiotensin-II infusion and in which PRA was very low. The ip. injection of an equimolar dose of SRIF completely reversed these effects of SRIF-A, but the administration of SRIF alone did not affect either PRA or PAC. Taken together, these data would suggest that, in the rat, endogenous SRIF exerts, under basal conditions, a two-fold maximum tonic inhibitory effect on both renin release by kidneys and aldosterone secretion by zona glomerulosa cells.
|
| 1354449 |
Vapreotide, a somatostatin analogue, in cryptosporidiosis and other AIDS-related diarrhoeal diseases |
10.1097/00002030-199207000-00015. |
AIDS |
Vapreotide, a somatostatin analogue, in cryptosporidiosis and other AIDS-related diarrhoeal diseases
Abstract
- To evaluate the efficacy and tolerance of vapreotide, a new somatostatin analogue, in the treatment of refractory AIDS-related diarrhoea.\n \n \n \n \n An open, non-comparative pilot trial.\n \n \n \n \n The trial was conducted in 10 medical centres in France.\n \n \n \n \n Thirty-four AIDS patients with chronic diarrhoea unresponsive to conventional antidiarrhoeal therapy were enrolled. Cryptosporidiosis was diagnosed in 21 out of 30 evaluable patients. Mean number of stools prior to therapy was 10.1 +/- 4.9 per day (range, 3-20 stools per day).\n \n \n \n \n After initial baseline studies, patients received subcutaneous vapreotide at escalating doses of 400 (23 patients) or 500 micrograms (seven patients), between two and six times daily.\n \n \n \n \n Efficacy was assessed after 14 days of therapy, when it was found to be effective. Responders were offered the opportunity to continue receiving therapy.\n \n \n \n \n Four patients demonstrated a complete response and 12 a partial response with greater than 50% reduction in daily stool emission. Fourteen patients did not respond to doses up to 2400 micrograms/day. Patients with conditions other than cryptosporidiosis had a significantly higher probability of response (P = 0.013), as did those with milder diarrhoea (less than 10 stools per day). Median duration of response was 1.5 months (range, 0.5-5 months); relapse occurred in five out of eight responders despite maintenance therapy. Toxicity was minimal.\n \n \n \n \n We conclude that AIDS patients with diarrhoea not caused by Cryptosporidium may benefit from vapreotide therapy.
|
| 1354985 |
Prothrombin Salakta: substitution of glutamic acid-466 by alanine reduces the fibrinogen clotting activity and the esterase activity |
10.1021/bi00148a005. |
Biochemistry |
Prothrombin Salakta: substitution of glutamic acid-466 by alanine reduces the fibrinogen clotting activity and the esterase activity
Abstract
- Structural studies on a hereditary abnormal prothrombin, prothrombin Salakta, have been performed to identify the difference responsible for its reduced fibrinogen clotting activity and its reduced esterase activity. Amino acid composition and sequence analyses of a peptide isolated from a lysylendopeptidase digest of the abnormal thrombin indicated that Glu-466 had been replaced by Ala. This amino acid substitution can result from a single nucleotide change in the codon for Glu-466 (GAG----GCG). The model building and the molecular dynamics simulation of thrombin Salakta suggest that the Glu-466----Ala substitution would change the proper conformation around the substrate binding site containing Trp-468, which is a unique surface loop on the thrombin molecule. This is the experimental and theoretical evidence supporting the role of the surface loop containing Trp-468 for the proper conformation of the substrate binding site.
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