Correlation between the effects of bombesin antagonists on cell proliferation and intracellular calcium concentration in Swiss 3T3 and HT-29 cell lines.

Bombesin (BN) acts as an autocrine mitogen in various human cancers. Several pseudononapeptide BN-(6-14) analogs with a reduced peptide bond between positions 13 and 14 have been shown to suppress the mitogenic activity of BN or gastrin-releasing peptide (GRP) when assessed by radioreceptor or proliferation assays and may have significant clinical applications. The search for potent and safe BN antagonists requires the evaluation of a large series of analogs in radioreceptor and proliferation assays. In this paper, we report that the ability of BN analogs to inhibit BN-induced calcium transients in Swiss 3T3 cells shows a high correlation with their inhibitory potency as evaluated by classical proliferation tests. The assay of calcium transients allows a rapid characterization of new BN analogs (in terms of minutes rather than days) and can be adapted as a labor and cost-effective screening step in the selection of potentially relevant BN antagonists for further characterization in cell proliferation systems. We also observed that results from the assay of calcium transients in Swiss 3T3 cells can be correlated with the results of the proliferative response in HT-29 cells, a cell line that does not seem to use the same early transmembrane ionic signal system. This result suggests that the calcium pathway is not mandatory for triggering cell division by the BN receptor.

Potent, structurally constrained agonists and competitive antagonists of corticotropin-releasing factor.

Predictive methods, physicochemical measurements, and structure activity relationship studies suggest that corticotropin-releasing factor (CRF; corticoliberin), its family members, and competitive antagonists (resulting from N-terminal deletions) usually assume an alpha-helical conformation when interacting with the CRF receptor(s). To test this hypothesis further, we have scanned the whole sequence of the CRF antagonist [D-Phe12,Nle21,38]r/hCRF-(12-41) (r/hCRF, rat/human CRF; Nle, norleucine) with an i-(i + 3) bridge consisting of the Glu-Xaa-Xaa-Lys scaffold. We have found astressin [cyclo(30-33)[D-Phe12,Nle21,38,Glu30,Lys33]r/ hCRF(12-41)] to be approximately 30 times more potent than [D-Phe12,Nle21,38]r/hCRF-(12-41), our present standard, and 300 times more potent than the corresponding linear analog in an in vitro pituitary cell culture assay. Astressin has low affinity for the CRF binding protein and high affinity (Ki = 2 nM) for the cloned pituitary receptor. Radioiodinated [D-125I-Tyr12]astressin was found to be a reliable ligand for binding assays. In vivo, astressin is significantly more potent than any previously tested antagonist in reducing hypophyseal corticotropin (ACTH) secretion in stressed or adrenalectomized rats. The cyclo(30-33)[Ac-Pro4,D-Phe12,Nle21,38,Glu30,Lys33++ +]r/hCRF-(4-41) agonist and its linear analog are nearly equipotent, while the antagonist astressin and its linear form vary greatly in their potencies. This suggests that the lactam cyclization reinstates a structural constraint in the antagonists that is normally induced by the N terminus of the agonist.

High-affinity neuropeptide Y receptor antagonists.

Neuropeptide Y (NPY) is one of the most abundant peptide transmitters in the mammalian brain. In the periphery it is costored and coreleased with norepinephrine from sympathetic nerve terminals. However, the physiological functions of this peptide remain unclear because of the absence of specific high-affinity receptor antagonists. Three potent NPY receptor antagonists were synthesized and tested for their biological activity in in vitro, ex vivo, and in vivo functional assays. We describe here the effects of these antagonists inhibiting specific radiolabeled NPY binding at Y1 and Y2 receptors and antagonizing the effects of NPY in human erythroleukemia cell intracellular calcium mobilization perfusion pressure in the isolated rat kidney, and mean arterial blood pressure in anesthetized rats.

Identification of receptor ligands and receptor subtypes using antagonists in a capillary electrophoresis single-cell biosensor separation system.

A capillary electrophoresis system with single-cell biosensors as a detector has been used to separate and identify ligands in complex biological samples. The power of this procedure was significantly increased by introducing antagonists that inhibited the cellular response from selected ligand-receptor interactions. The single-cell biosensor was based on the ligand-receptor binding and G-protein-mediated signal transduction pathways in PC12 and NG108-15 cell lines. Receptor activation was measured as increases in cytosolic free calcium ion concentration by using fluorescence microscopy with the intracellular calcium ion indicator fluo-3-acetoxymethyl ester. Specifically, a mixture of bradykinin (BK) and acetylcholine (ACh) was fractionated and the components were identified by inhibiting the cellular response with icatibant (HOE 140), a selective antagonist to the BK B2 receptor subtype (B2BK), and atropine, an antagonist to muscarinic ACh receptor subtypes. Structurally related forms of BK were also identified based on inhibiting B2BK receptors. Applications of this technique include identification of endogenous BK in a lysate of human hepatocellular carcinoma cells (Hep G2) and screening for bioactivity of BK degradation products in human blood plasma. The data demonstrate that the use of antagonists with a single-cell biosensor separation system aids identification of separated components and receptor subtypes.

FDA narcotic treatment programs directory.

Mode of access: Internet.

The Antagonists,

The first of a trilogy completed by his Richard Furlong and Achievement.

The superhuman antagonists, and other poems,

The "fundamental idea" of "The superhuman antagonists" (p. 1-50) is borrowed from "ancient Persian mythology." cf. Pref.

An essay on alcoholic & narcotic substances, as articles of common use : addressed particularly to students /

Second edition issued in 1830 under title: An essay on temperance, addressed particularly to students ...

Treasury decisions under the customs, internal revenue, industrial alcohol, narcotic and other laws.

Title varies slightly.

Revocation or denial of seamen's documents to narcotic law violators. Hearing before a subcommittee of the Committee on Interstate and Foreign Commerce, United States Senate, Eighty-third Congress, second session, on H. R. 8538, an act to provide for the revocation or denial of Merchant Marine documents to persons involved in certain narcotics violations. June 16, 1954.

Mode of access: Internet.

Narcotic Act

Mode of access: Internet.

Comprehensive narcotic addiction and drug abuse care and control act of 1969. Hearings, Ninety-first Congress, first and second sessions.

Hearings held Sept. 18-Nov. 3, 1969, in Washington, D.C.; Jan. 26, 1970, in Cherry Hill, N.J.

Summary statistics, civil commitment program for narcotic addicts

Mode of access: Internet.

Compilation of selected public health laws: including particularly the Public health service act, the Clean air act, the Solid waste disposal act, the Mental retardation facilities and community mental health centers construction act, and the Narcotic addict rehabilitation act, the Federal food, drug, and cosmetic act, the Federal hazardous substances act, and the fair packaging and labeling act.

Prepared for the use of the Senate Committee on Labor and Public Welfare and the House Committee on Interstate and Foreign Commerce.

Potent cyclic antagonists of the complement C5a receptor on human polymorphonulcear leukocytes. Relationships between structures and activity

Human C5a is a plasma protein with potent chemoattractant and pro-inflammatory properties, and its overexpression correlates with severity of inflammatory diseases. C5a binds to its G protein-coupled receptor (C5aR) on polymorphonuclear leukocytes (PMNLs) through a high-affinity helical bundle and a low-affinity C terminus, the latter being solely responsible for receptor activation. Potent and selective C5a antagonists are predicted to be effective anti-inflammatory drugs, but no pharmacophore for small molecule antagonists has yet been developed, and it would significantly aid drug design. We have hypothesized that a turn conformation is important for activity of the C terminus of C5a and herein report small cyclic peptides that are stable turn mimics with potent antagonism at C5aR on human PMNLs. A comparison of solution structures for the C terminus of C5a, small acyclic peptide ligands, and cyclic antagonists supports the importance of a turn for receptor binding. Competition between a cyclic antagonist and either C5a or an acyclic agonist for C5aR on PMNLs supports a common or overlapping binding site on the C5aR. Structure-activity relationships for 60 cyclic analogs were evaluated by competitive radioligand binding with C5a (affinity) and myeloperoxidase release (antagonist potency) from human PMNLs, with 20 compounds having high antagonist potencies (IC50, 20 nM(-1) muM). Computer modeling comparisons reveal that potent antagonists share a common cyclic backbone shape, with affinity-determining side chains of defined volume projecting from the cyclic scaffold. These results define a new pharmacophore for C5a antagonist development and advance our understanding of ligand recognition and receptor activation of this G protein-coupled receptor.