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T L Peeters - One of the best experts on this subject based on the ideXlab platform.

  • Contractile effects and intracellular Ca2+ signalling induced by motilin and erythromycin in the circular smooth muscle of human colon
    Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society, 2001
    Co-Authors: G Van Assche, Inge Depoortere, Theo Thijs, Jozef Janssens, Ludwig Missiaen, Freddy Penninckx, H. Takanashi, K. Geboes, T L Peeters
    Abstract:

    Motilin has excitatory effects on the colon of the rabbit and the dog, but little is known of its effect on the human colon. The aim of this study was to investigate the effects induced by motilin and erythromycin A (EMA) on muscle strips and on single cells from primary cultures from human colon. Isotonic contraction was recorded in circular muscle strips from macroscopically normal resection specimens of patients operated on for colonic neoplasm. Agonist-induced intracellular Ca2+ ([Ca2+]i) signalling was studied in primary cultures of colonic smooth-muscle cells using the ratiometric Ca2+ indicator Indo 1, on a laser-scanning confocal epifluorescence microscope. In circular muscle strips, norleucine13-porcine motilin ([Nle13]-pm)and EMA induced tonic contractions with an EC50 of 92 +/- 21 nmol L(-1) and 31 +/- 16 micromol L(-1), respectively. The maximal contraction was 21 +/- 4% (motilin) and 33 +/- 12% (EMA) of the response to 10(-4) mol L(-1) acetylcholine (ACh). The motilin antagonist OHM-11526 (10(-5.5) mol L(-1)) abolished the effects of both [Nle13]-pm and EMA. Neither tetrodotoxin (10(-5.5) mol L(-1)), L-nitro-D-arginine methyl ester (L-NAME) (10(-3.5) mol L(-1)) nor guanethidine (10(-5) mol L(-1)) interfered with the effects of [Nle13]-pm or EMA. [Nle13]-pm (10(-11)-10(-6) mol L(-1)) induced rises of [Ca2+]i in cultured colonic myocytes. At 10(-6) mol L-1, 94% of the cells responded, and half of the cells responded at 1.4 nmol L(-1) [Nle13]-pm. 81% (35/43) and 95% (75/79) responded to EMA (10(-6) mol L(-1)) and acetylcholine (ACh, 10(-4) mol L(-1)), respectively. The motilin antagonist GM-109 inhibited motilin- and EMA-induced [Ca2+]i rises. In the absence of extracellular Ca2+, only 13% (7/52) of the cells responded to [Nle13]-pm (10(-6) mol L(-1)) vs. 90% (47/52) to ACh (10(-4) mol L(-1)). Motilin and EMA have direct excitatory effects on circular smooth muscle from the human colon and these effects are mediated via a smooth-muscle motilin receptor. These findings suggest that motilin may regulate colonic motility and that Motilides may have therapeutic potential for the treatment of colonic hypomotility.

  • Effect of different prokinetic agents and a novel enterokinetic agent on postoperative ileus in rats.
    Gut, 1999
    Co-Authors: B.y. De Winter, T L Peeters, Guy E. Boeckxstaens, J. G. De Man, Tom G. Moreels, J.a.j. Schuurkes, A. G. Herman, P. A. Pelckmans
    Abstract:

    Background/Aim—The eVects of diVerent prokinetic agents, the Motilide erythromycin and the substituted benzamides metoclopramide and cisapride, were investigated in a rat model of postoperative ileus. These eVects were compared with that of granisetron, a 5-hydroxytryptamine (5-HT3) receptor antagonist, and a novel enterokinetic agent, prucalopride, a 5-HT4 receptor agonist. Methods—DiVerent degrees of inhibition of gastrointestinal transit, measured by the migration of Evans blue, were achieved by skin incision, laparotomy, or laparotomy plus mechanical stimulation of the gut. Results—Metoclopramide decreased the transit after laparotomy with or without mechanical stimulation, whereas cisapride increased it after all three operations. Granisetron had no eVect on the transit after the three operations when given alone. Prucalopride tended to increase the transit after laparotomy with or without mechanical stimulation when given alone. However, statistical significance was only reached when prucalopride was combined with granisetron. Erythromycin, a motilin receptor agonist, did not improve postoperative ileus in the rat. Conclusions—Cisapride, but not metoclopramide or erythromycin, is able to improve postoperative ileus in the rat.The results suggest that a combination of 5-HT3 receptor antagonist and 5-HT4 receptor agonist properties may be required to obtain a beneficial eVect on surgery induced ileus in the rat. Furthermore, they indirectly indicate that stimulation of the excitatory mechanisms is not able to overcome the inhibitory influence of the neural reflex pathways activated during abdominal surgery. (Gut 1999;45:713‐718)

  • Potential of Motilides in the Treatment of Hypomotility Syndromes
    Problems of the Gastrointestinal Tract in Anesthesia the Perioperative Period and Intensive Care, 1999
    Co-Authors: T L Peeters
    Abstract:

    The 22-amino-acid polypeptide motilin was discovered as a result of the study of the relationship between changes in duodenal pH and gastric motility (Brown et al. 1966, 1972). As soon as the synthesis of the peptide was achieved, it was logical for the effect on gastric emptying to be explored. However, the results were contradictory: acceleration, lack of effect and retardation were all reported, and given that the peptide was expensive and the methodology for measuring gastric emptying tedious, the topic was abandoned. Because intestinal pH changes also affect pressure in the lower esophageal sphincter (LES), attention was then directed to the effect of motilin on the LES, and the induction of a rise in basal LES pressure was soon demonstrated. Then the study of the physiology of motilin took a new direction with the discovery of the relation between plasma motilin levels and phase 3 of the migrating motor complex (MMC) in dogs (Itoh et al. 1978), just a few years after the MMC had been described in this species. This relation was soon confirmed in humans, and the finding that in dogs and in humans exogenous motilin induced phase 3 activity suggested that motilin’s physiological role was the regulation of interdigestive motor activity. Despite numerous studies this hypothesis remains controversial.

  • Concentration-dependent stimulation of cholinergic motor nerves or smooth muscle by [Nle13]motilin in the isolated rabbit gastric antrum.
    European journal of pharmacology, 1997
    Co-Authors: G Van Assche, I. Depoortere, T Thijs, J J Janssens, T L Peeters
    Abstract:

    In man, rabbit and cat, the effects of motilin and Motilides are neurally mediated in vivo, whereas in vitro binding and contractility studies suggest the presence of a smooth muscular receptor. The aim of this study was to investigate in vitro interactions of motilin with the enteric excitatory neurotransmission in the gastric antrum of the rabbit. Circular muscle strips from the pre-pyloric antrum were subjected to electrical field stimulation (1 ms, 1-32 Hz, 10 s train) and muscle twitch responses were recorded isometrically. Induced twitch responses were frequency dependent (1-32 Hz) and entirely neurogenic (tetrodotoxin sensitive). [Nle13]motilin dose-dependently (10[-9]-10[-8] M) enhanced the amplitude of, atropine sensitive, evoked contractions. At 4 Hz the response, expressed as a % of the response to 32 Hz, increased from 15.5 +/- 4.1% (control) to 28.1 +/- 5.8% (motilin 10[-9] M), and to 45.8 +/- 3.6% (motilin 10[-8.5] M) (P < 0.05). This effect was not inhibited by hexamethonium (10[-3.3] M) but was abolished by the motilin receptor antagonist GM-109 (10[-5] M). In unstimulated strips, motilin induced phasic-tonic contractions with a threshold concentration of 10[-8] M and an pEC50 of 7.48, which were also inhibited by GM-109 (10[-5] M) but not by tetrodotoxin (10[-5.5] M). The maximal tension, frequency and dose-dependency of carbachol-induced contractions were not influenced by motilin (pEC50, carbachol: 6.48 +/- 0.06 (control), 6.49 +/- 0.07 (motilin)). In conclusion, motilin enhances contractions induced by electrical field stimulation in the rabbit antrum by a post-ganglionic interaction with the cholinergic neurotransmission in vitro at low doses and interacts directly with antral smooth muscle at high doses. This model is an accurate reflection of the in vivo effects of motilin and provides a tool to study neurogenic and myogenic actions of motilin and Motilides in vitro.

  • Concentration-dependent stimulation of cholinergic motor nerves or smooth muscle by [Nle13]motilin in the isolated rabbit gastric antrum
    European Journal of Pharmacology, 1997
    Co-Authors: G Vanassche, Inge Depoortere, Theo Thijs, Jozef Janssens, T L Peeters
    Abstract:

    In man, rabbit and cat, the effects of motilin and Motilides are neurally mediated in vivo, whereas in vitro binding and contractility studies suggest the presence of a smooth muscular receptor. The aim of this study was to investigate in vitro interactions of motilin with the enteric excitatory neurotransmission in the gastric antrum of the rabbit. Circular muscle strips from the pre-pyloric antrum were subjected to electrical field stimulation (1 ms, 1-32 Hz, 10 s train) and muscle twitch responses were recorded isometrically. Induced twitch responses were frequency dependent (1-32 Hz) and entirely neurogenic (tetrodotoxin sensitive). [Nle(13)]motilin dose-dependently (10(-9)-10(-8) M) enhanced the amplitude of, atropine sensitive, evoked contractions. At 4 Hz the response, expressed as a % of the response to 32 Hz, increased from 15.5 +/- 4.1% (control) to 28.1 +/- 5.8% (motiiin 10(-9) M), and to 45.8 +/- 3.6% (motilin 10(-8.5) M) (P < 0.05). This effect was not inhibited by hexamethonium (10(-3.3) M) but was abolished by the motilin receptor antagonist GM-109 (10(-5) M). In unstimulated strips, motilin induced phasic-tonic contractions with a threshold concentration of 10(-8) M and an pEC(50) of 7.48, which were also inhibited by GM-109 (10(-5) M) but not by tetrodotoxin (10(-5.5) M). The maximal tension, frequency and dose-dependency of carbachol-induced contractions were not influenced by motilin (pEC(50), carbachol: 6.48 +/- 0.06 (control), 6.49 +/- 0.07 (motilin)). In conclusion, motilin enhances contractions induced by electrical field stimulation in the rabbit antrum by a post-ganglionic interaction with the cholinergic neurotransmission in vitro at low doses and interacts directly with antral smooth muscle at high doses. This model is an accurate reflection of the in vivo effects of motilin and provides a tool to study neurogenic and myogenic actions of motilin and Motilides in vitro. (C) 1997 Elsevier Science B.V.

Theo L. Peeters - One of the best experts on this subject based on the ideXlab platform.

  • Structure-Activity Relation of Erythromycin-Related Macrolides in Inducing Contractions and in Displacing Bound Motilin in Rabbit Duodenum
    Neurogastroenterology & Motility, 2008
    Co-Authors: Inge Depoortere, Theo L. Peeters, Gert Matthijs, T Cachet, Jos Hoogmartens, Gaston Vantrappen
    Abstract:

    ERYTHROMYCIN A appears to be a motilin agonist, although motilin (a peptide) and erythromycin A (a 14-member macrolide antibiotic carrying two sugar substituents) are structurally totally unrelated. To obtain information concerning the structural requirements for erythromycin's interaction with the motilin receptor, we studied 60 derivatives and seven fragments of erythromycin, and six macrolides with a 16-member ring structure. For every compound the ability to displace motilin bound to a crude homogenate of rabbit antral smooth muscle tissue and the ability to induce contractions in rabbit duodenal segments were determined. For both parameters dose-response curves were obtained, and the negative logarithms of the concentrations inhibiting 50% of the binding (pIC-50) or inducing 50% of the maximum response to a maximal contractile dose of acetylcholine (pEC-50) were determined. All macrolides with a 16-member ring structure were inactive in both types of experiments. In all erythromycin derivatives variations of the ring structure had a marked effect. For both parameters the order of potency was enol ether > pseudo–enol ether > parent ring > pseudo-hemiketal > anhydro. The two sugars attached to the ring were important too, because removing either or both of them resulted in an inactive compound. Modifications of the dimethylamino group of the desosamine sugar affect the potency, while the orientation of the cladinose moiety seems to be important too. For all active compounds, both parameters studied were well correlated (r= 0.80, p < 0.001). Our results support the concept that erythomycin-like macrolides are motilin agonists. The structural requirements of these “Motilides” involve the ring structure, especially the part that can be transformed into an enol ether, and both attached sugars. Potent derivatives may prove to be useful as gastrokinetic agents with a very specific target zone.

  • Differences in motilin receptor desensitization after stimulation with motilin or Motilides are due to alternative receptor trafficking
    Biochemical pharmacology, 2007
    Co-Authors: Anna Mitselos, Theo L. Peeters, Pieter Vanden Berghe, Inge Depoortere
    Abstract:

    Abstract Backgrounds & aims The motilin receptor (MTLR) is an important therapeutic target for treatment of hypomotility disorders. The negative outcome in clinical trials with the motilin agonist, ABT-229, indicated that desensitization may limit the therapeutic usefulness of Motilides. We therefore compared the mechanisms involved in the intracellular trafficking of the MTLR after stimulation with motilin, erythromycin-A (EM-A) or ABT-229. Methods Desensitization was studied by measuring changes in Ca 2+ rises and by receptor binding studies in CHO cells co-expressing the Ca 2+ indicator apoaequorin and the MTLR, C-terminally tagged with EGFP. Receptor phosphorylation was studied by immunoprecipitation. MTLR-EGFP trafficking to organelles and translocation of β-arrestins were visualized by fluorescence microscopy. Results Agonist-induced desensitization of the MTLR was due to receptor internalization with potencies (p-int 50 ) in the order of: ABT-229 (8.3) > motilin (7.86) > EM-A (4.77) but with no differences in the internalization kinetics ( t 1/2 : ∼25 min). The percentage cell surface receptor loss was more profound after exposure to ABT-229 (88 ± 1%) than to motilin (63 ± 10%) or EM-A (34 ± 2%). For motilin and EM-A MTLR phosphorylation probably occurs via G protein-coupled receptor kinases while for ABT-229 phosphorylation was also protein kinase C dependent. All agonists translocated cytosolic β-arrestin-2 with greater affinity to the plasma membrane than β-arrestin-1. After internalization the MTLR co-localized with transferrin but not with cathepsin D. After stimulation with motilin and EM-A the t 1/2 for MTLR resensitization was 3 h and 1 h, respectively but amounted 26 h for ABT-229. Conclusion Our results suggest that the resensitization kinetics determine the desensitization properties of the motilin agonists.

  • Potential of ghrelin as a therapeutic approach for gastrointestinal motility disorders.
    Current opinion in pharmacology, 2006
    Co-Authors: Theo L. Peeters
    Abstract:

    Ghrelin was first discovered as a peptide involved in growth hormone release, but has now emerged as a new player in the regulation of gastrointestinal function. Ghrelin is structurally and functionally related to motilin. Like motilin, it induces a specific motor pattern in the fasted state and acts postprandially to accelerate gastric emptying. There is no apparent cross-reactivity with motilin at the receptor level. Ghrelin agonists have the same potential as motilin agonists, and applications in post-operative ileus and gastroparesis have already been explored. Although promising, there is still the need to avoid side effects and the problems encountered with Motilides. This will require drugs with an appropriate pharmacokinetic profile. In addition, the dosage regimen and target population should be carefully taken into consideration when planning clinical trials.

  • Desensitization of the Human Motilin Receptor by Motilides
    The Journal of pharmacology and experimental therapeutics, 2005
    Co-Authors: Leen Thielemans, Christopher Carreras, Inge Depoortere, Jason Perret, Patrick Robberecht, Yaoquan Liu, Theo Thijs, Emmanuel Burgeon, Theo L. Peeters
    Abstract:

    Tachyphylaxis may have contributed to the failure of the Motilide ABT-229 [ N -ethyl, N -methyl 4′′ deoxy erythromycin (EM)-B enolether] in clinical trials. We compared the desensitizing potency of structurally related Motilides [EM-A, EM-A enolether (ME4), N -ethyl, N -methyl EM-A (ME36), EM-B enolether (ME67), N -ethyl, N -methyl EM-A enolether (EM523), ABT-229 and 4′′ deoxy EM-A enolether (KOS1326)] in a Chinese hamster ovary (CHO)-K1 cell line expressing the human motilin receptor (MTLR) and in rabbit duodenal segments. CHO-MTLR cells were preincubated with Motilides prior to stimulation with motilin. The negative logarithm of the preincubation concentration reducing the maximal motilin-induced Ca 2+ flux to 50% was calculated (pDC 50 ). Internalization was visualized in CHO-K1 cells containing an enhanced green fluorescent protein (EGFP)-tagged MTLR and quantified in binding experiments. The contractile response of repeated stimulations was measured in duodenal segments. In CHO-MTLR cells, the pDC 50 was ABT-229 (8.78) > motilin (7.77) > EM-A (4.78), different from their order of potency to induce Ca 2+ release (pEC 50 ): motilin (9.39) > ABT-229 (8.46) > EM-A (7.11). In cells with the EGFP-tagged MTLR, ABT-229 decreased membrane fluorescence by 25 ± 2% compared with 16 ± 2% for motilin and 8 ± 2% for EM-A. Binding studies confirmed that EM-A did not induce MTLR internalization (residual binding 96 ± 4% compared with motilin, 31 ± 3% and ABT-229, 21 ± 1%). Comparison of the pDC 50 and pEC 50 values of the other Motilides ME4 (5.90; 8.08), ME67 (6.03; 8.12), ME36 (3.32; 6.62), EM-523 (6.02; 8.22), and KOS1326 (7.32; 8.14) suggested that the strong desensitizing properties of ABT-229 are mostly related to the removal of the 4″-OH of the cladinose sugar. The decline of the contractile response in duodenal segments correlated with the pDC 50 . The ability to desensitize and internalize the MTLR is not only determined by potency. This may be an important criterion for the development of a clinically useful compound.

  • The motilin pharmacophore in CHO cells expressing the human motilin receptor.
    Biochemical and biophysical research communications, 2002
    Co-Authors: Leen Thielemans, Inge Depoortere, Jozef Vanden Broeck, Theo L. Peeters
    Abstract:

    Abstract We performed a structure–activity study with the human motilin receptor, which was recently cloned from thyroid tissue. N-terminal fragments, Ala-analogs of motilin, and Motilides were tested in a cell line that expresses the cloned human motilin receptor and apoaequorin. Full potency to induce calcium fluxes was obtained with N-terminal fragments of 14 amino acids. Motilin fragments 1–14 in which residues 1 (Phe), 4 (Ile), and 7 (Tyr) were replaced by Ala showed the largest reduction in potency. Only Motilides with an enol configuration had markedly higher potencies compared to erythromycin A. The potencies to induce Ca 2+ fluxes correlated strongly with rabbit binding and contractility data, suggesting that the cloned receptor is indeed the motilin receptor, responsible for contractile effects. Conservation of the motilin pharmacophore in evolution indicates an important physiological role of motilin.

Gaston Vantrappen - One of the best experts on this subject based on the ideXlab platform.

  • Structure-Activity Relation of Erythromycin-Related Macrolides in Inducing Contractions and in Displacing Bound Motilin in Rabbit Duodenum
    Neurogastroenterology & Motility, 2008
    Co-Authors: Inge Depoortere, Theo L. Peeters, Gert Matthijs, T Cachet, Jos Hoogmartens, Gaston Vantrappen
    Abstract:

    ERYTHROMYCIN A appears to be a motilin agonist, although motilin (a peptide) and erythromycin A (a 14-member macrolide antibiotic carrying two sugar substituents) are structurally totally unrelated. To obtain information concerning the structural requirements for erythromycin's interaction with the motilin receptor, we studied 60 derivatives and seven fragments of erythromycin, and six macrolides with a 16-member ring structure. For every compound the ability to displace motilin bound to a crude homogenate of rabbit antral smooth muscle tissue and the ability to induce contractions in rabbit duodenal segments were determined. For both parameters dose-response curves were obtained, and the negative logarithms of the concentrations inhibiting 50% of the binding (pIC-50) or inducing 50% of the maximum response to a maximal contractile dose of acetylcholine (pEC-50) were determined. All macrolides with a 16-member ring structure were inactive in both types of experiments. In all erythromycin derivatives variations of the ring structure had a marked effect. For both parameters the order of potency was enol ether > pseudo–enol ether > parent ring > pseudo-hemiketal > anhydro. The two sugars attached to the ring were important too, because removing either or both of them resulted in an inactive compound. Modifications of the dimethylamino group of the desosamine sugar affect the potency, while the orientation of the cladinose moiety seems to be important too. For all active compounds, both parameters studied were well correlated (r= 0.80, p < 0.001). Our results support the concept that erythomycin-like macrolides are motilin agonists. The structural requirements of these “Motilides” involve the ring structure, especially the part that can be transformed into an enol ether, and both attached sugars. Potent derivatives may prove to be useful as gastrokinetic agents with a very specific target zone.

  • Distribution and characterization of motilin receptors in the cat.
    Peptides, 1993
    Co-Authors: Inge Depoortere, Theo L. Peeters, Gaston Vantrappen
    Abstract:

    Abstract We demonstrate binding of [ 125 I][Nle 13 -po]motilin to homogenates of cat gastric and small intestinal, but not to colonic smooth muscle tissue. The density was ( B max in fmol/mg protein): 0 (fundus); 12 ± 2 (corpus); 22 ± 3 (antrum); 55 ± 12 (duodenum); 44 ± 10 (jejunum); 17 ± 1 (ileum); 0 (colon). A significant ( p ) difference was found between the dissociation constant for motilin in the stomach ( pK d = 8.84 ± 0.06 ) and in the small intestine ( pK d = 8.58 ± 0.08 ). The Motilides erythromycin-A (EM-A), EM-523, and EM-A N- oxide displaced labeled [Nle 13 -po]motilin bound to cat duodenal receptor with potencies ( pK d ) of 5.47 ± 0.23 , 7.60 ± 0.24 , and 13 -po]motilin fragments showed that the N- terminus of motilin interacts with the receptor. In the tissue bath, duodenal strips mounted in the longitudinal direction responded to motilin, EM-523, and EM-A (pEC 50 : 8.29 ± 0.08 ; 7.12 ± 0.12 ; 5.99 ± 0.15 ). The compounds had a comparable intrinsic activity ( 83 ± 3% ; 80 ± 5% ; 82 ± 5% of the response to ACh), which was unaffected by atropine, TTX, hexamethonium, and zacopride but reduced by verapamil and calcium-free medium. Cat stomach and small intestine possess smooth muscle motilin receptors, which have comparable properties as those found in man and in rabbit.

  • Ca^2+ dependence of Motilide-induced contractions in rabbit duodenal muscle strips in vitro
    Naunyn-Schmiedeberg's Archives of Pharmacology, 1991
    Co-Authors: T L Peeters, Gert Matthijs, Gaston Vantrappen
    Abstract:

    Recent studies suggested that certain erythromycin A (EM-A) derivatives are motilin receptor agonists. As proposed by Itoh they may be called “Motilides”. We have investigated the Ca^2+-dependence of contractions induced by two potent Motilides, ME-34 [de( N -methyl) 8,9-anhydroeryhtromycin A 6,9-hemiacetal] and EM-523 [de( N -methyl)- N -ethyl-8,9-anhydro-erythromycin A 6,9-hemiacetal], in duodenal tissues and compared the results with those previously obtained with motilin. Isometric and isotonic contractile responses of isolated longitudinal muscle sheets from the rabbit duodenum were tested under normal, Ca^2+-free and depolarizing conditions. Prior to stimulation with Motilides, the maximal response to acetylcholine was recorded and all responses were always expressed as a percentage of this response. Both Motilides induced contractions in normally polarized tissue, with an EC_50 of 26 ± 5 nM for ME-34 ( n = 7), and 27 ± 5 nM for EM-5231 ( n = 16) and maximal responses of respectively 88 ± 4% and 80 ± 3%. Like motilin, both compounds induced an ‘extra’-contraction in depolarized tissues. The EM-523 response in 140 mM K^+under isotonic conditions was 84 ± 3% ( n = 5) at 10^−5 M , with an EC_50 that was shifted to 65 ± 18 nM. Similar figures were obtained for ME-34. When Ca^2+ was added to Ca^2+-depleted strips, half-maximal Ca^2+ values (in mM) were 1.10 ± 0.11 ( n = 9) for EM-523 and 1.13 ± 0.12 ( n = 3) for ME-34, as compared with 1.12 ± 0.13 ( n = 7) for motilin and 2.8 ± 1.1 ( n = 9) for K^+. Both ME-34 and EM-523 also induced a transient contraction in Ca^+-free solutions under isometric conditions. The response to EM-523 (5 × 10^−6 M ) was 49 ± 15% ( n = 4) after 3 min. A maximal EM-523 -stimulation reduced a subsequent ACh response by 78 ± 7%, whereas EM-523 and ME-34 could not induce a contraction after ACh. We conclude that Motilides depend upon external Ca^2+ to a similar extent to motilin. Like motilin, they are also able to mobilize intracellular Ca Z + stores. Our results support the hypothesis that Motilides act on motilin receptors.

  • Ca2+ dependence of Motilide-induced contractions in rabbit duodenal muscle strips in vitro.
    Naunyn-Schmiedeberg's archives of pharmacology, 1991
    Co-Authors: T L Peeters, Gert Matthijs, Gaston Vantrappen
    Abstract:

    Recent studies suggested that certain erythromycin A (EM-A) derivatives are motilin receptor agonists. As proposed by Itoh they may be called “Motilides”. We have investigated the Ca2+-dependence of contractions induced by two potent Motilides, ME-34 [de(N-methyl) 8,9-anhydroeryhtromycin A 6,9-hemiacetal] and EM-523 [de(N-methyl)-N-ethyl-8,9-anhydro-erythromycin A 6,9-hemiacetal], in duodenal tissues and compared the results with those previously obtained with motilin.

Inge Depoortere - One of the best experts on this subject based on the ideXlab platform.

  • Structure-Activity Relation of Erythromycin-Related Macrolides in Inducing Contractions and in Displacing Bound Motilin in Rabbit Duodenum
    Neurogastroenterology & Motility, 2008
    Co-Authors: Inge Depoortere, Theo L. Peeters, Gert Matthijs, T Cachet, Jos Hoogmartens, Gaston Vantrappen
    Abstract:

    ERYTHROMYCIN A appears to be a motilin agonist, although motilin (a peptide) and erythromycin A (a 14-member macrolide antibiotic carrying two sugar substituents) are structurally totally unrelated. To obtain information concerning the structural requirements for erythromycin's interaction with the motilin receptor, we studied 60 derivatives and seven fragments of erythromycin, and six macrolides with a 16-member ring structure. For every compound the ability to displace motilin bound to a crude homogenate of rabbit antral smooth muscle tissue and the ability to induce contractions in rabbit duodenal segments were determined. For both parameters dose-response curves were obtained, and the negative logarithms of the concentrations inhibiting 50% of the binding (pIC-50) or inducing 50% of the maximum response to a maximal contractile dose of acetylcholine (pEC-50) were determined. All macrolides with a 16-member ring structure were inactive in both types of experiments. In all erythromycin derivatives variations of the ring structure had a marked effect. For both parameters the order of potency was enol ether > pseudo–enol ether > parent ring > pseudo-hemiketal > anhydro. The two sugars attached to the ring were important too, because removing either or both of them resulted in an inactive compound. Modifications of the dimethylamino group of the desosamine sugar affect the potency, while the orientation of the cladinose moiety seems to be important too. For all active compounds, both parameters studied were well correlated (r= 0.80, p < 0.001). Our results support the concept that erythomycin-like macrolides are motilin agonists. The structural requirements of these “Motilides” involve the ring structure, especially the part that can be transformed into an enol ether, and both attached sugars. Potent derivatives may prove to be useful as gastrokinetic agents with a very specific target zone.

  • Differences in motilin receptor desensitization after stimulation with motilin or Motilides are due to alternative receptor trafficking
    Biochemical pharmacology, 2007
    Co-Authors: Anna Mitselos, Theo L. Peeters, Pieter Vanden Berghe, Inge Depoortere
    Abstract:

    Abstract Backgrounds & aims The motilin receptor (MTLR) is an important therapeutic target for treatment of hypomotility disorders. The negative outcome in clinical trials with the motilin agonist, ABT-229, indicated that desensitization may limit the therapeutic usefulness of Motilides. We therefore compared the mechanisms involved in the intracellular trafficking of the MTLR after stimulation with motilin, erythromycin-A (EM-A) or ABT-229. Methods Desensitization was studied by measuring changes in Ca 2+ rises and by receptor binding studies in CHO cells co-expressing the Ca 2+ indicator apoaequorin and the MTLR, C-terminally tagged with EGFP. Receptor phosphorylation was studied by immunoprecipitation. MTLR-EGFP trafficking to organelles and translocation of β-arrestins were visualized by fluorescence microscopy. Results Agonist-induced desensitization of the MTLR was due to receptor internalization with potencies (p-int 50 ) in the order of: ABT-229 (8.3) > motilin (7.86) > EM-A (4.77) but with no differences in the internalization kinetics ( t 1/2 : ∼25 min). The percentage cell surface receptor loss was more profound after exposure to ABT-229 (88 ± 1%) than to motilin (63 ± 10%) or EM-A (34 ± 2%). For motilin and EM-A MTLR phosphorylation probably occurs via G protein-coupled receptor kinases while for ABT-229 phosphorylation was also protein kinase C dependent. All agonists translocated cytosolic β-arrestin-2 with greater affinity to the plasma membrane than β-arrestin-1. After internalization the MTLR co-localized with transferrin but not with cathepsin D. After stimulation with motilin and EM-A the t 1/2 for MTLR resensitization was 3 h and 1 h, respectively but amounted 26 h for ABT-229. Conclusion Our results suggest that the resensitization kinetics determine the desensitization properties of the motilin agonists.

  • Desensitization of the Human Motilin Receptor by Motilides
    The Journal of pharmacology and experimental therapeutics, 2005
    Co-Authors: Leen Thielemans, Christopher Carreras, Inge Depoortere, Jason Perret, Patrick Robberecht, Yaoquan Liu, Theo Thijs, Emmanuel Burgeon, Theo L. Peeters
    Abstract:

    Tachyphylaxis may have contributed to the failure of the Motilide ABT-229 [ N -ethyl, N -methyl 4′′ deoxy erythromycin (EM)-B enolether] in clinical trials. We compared the desensitizing potency of structurally related Motilides [EM-A, EM-A enolether (ME4), N -ethyl, N -methyl EM-A (ME36), EM-B enolether (ME67), N -ethyl, N -methyl EM-A enolether (EM523), ABT-229 and 4′′ deoxy EM-A enolether (KOS1326)] in a Chinese hamster ovary (CHO)-K1 cell line expressing the human motilin receptor (MTLR) and in rabbit duodenal segments. CHO-MTLR cells were preincubated with Motilides prior to stimulation with motilin. The negative logarithm of the preincubation concentration reducing the maximal motilin-induced Ca 2+ flux to 50% was calculated (pDC 50 ). Internalization was visualized in CHO-K1 cells containing an enhanced green fluorescent protein (EGFP)-tagged MTLR and quantified in binding experiments. The contractile response of repeated stimulations was measured in duodenal segments. In CHO-MTLR cells, the pDC 50 was ABT-229 (8.78) > motilin (7.77) > EM-A (4.78), different from their order of potency to induce Ca 2+ release (pEC 50 ): motilin (9.39) > ABT-229 (8.46) > EM-A (7.11). In cells with the EGFP-tagged MTLR, ABT-229 decreased membrane fluorescence by 25 ± 2% compared with 16 ± 2% for motilin and 8 ± 2% for EM-A. Binding studies confirmed that EM-A did not induce MTLR internalization (residual binding 96 ± 4% compared with motilin, 31 ± 3% and ABT-229, 21 ± 1%). Comparison of the pDC 50 and pEC 50 values of the other Motilides ME4 (5.90; 8.08), ME67 (6.03; 8.12), ME36 (3.32; 6.62), EM-523 (6.02; 8.22), and KOS1326 (7.32; 8.14) suggested that the strong desensitizing properties of ABT-229 are mostly related to the removal of the 4″-OH of the cladinose sugar. The decline of the contractile response in duodenal segments correlated with the pDC 50 . The ability to desensitize and internalize the MTLR is not only determined by potency. This may be an important criterion for the development of a clinically useful compound.

  • The motilin pharmacophore in CHO cells expressing the human motilin receptor.
    Biochemical and biophysical research communications, 2002
    Co-Authors: Leen Thielemans, Inge Depoortere, Jozef Vanden Broeck, Theo L. Peeters
    Abstract:

    Abstract We performed a structure–activity study with the human motilin receptor, which was recently cloned from thyroid tissue. N-terminal fragments, Ala-analogs of motilin, and Motilides were tested in a cell line that expresses the cloned human motilin receptor and apoaequorin. Full potency to induce calcium fluxes was obtained with N-terminal fragments of 14 amino acids. Motilin fragments 1–14 in which residues 1 (Phe), 4 (Ile), and 7 (Tyr) were replaced by Ala showed the largest reduction in potency. Only Motilides with an enol configuration had markedly higher potencies compared to erythromycin A. The potencies to induce Ca 2+ fluxes correlated strongly with rabbit binding and contractility data, suggesting that the cloned receptor is indeed the motilin receptor, responsible for contractile effects. Conservation of the motilin pharmacophore in evolution indicates an important physiological role of motilin.

  • Contractile effects and intracellular Ca2+ signalling induced by motilin and erythromycin in the circular smooth muscle of human colon
    Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society, 2001
    Co-Authors: G Van Assche, Inge Depoortere, Theo Thijs, Jozef Janssens, Ludwig Missiaen, Freddy Penninckx, H. Takanashi, K. Geboes, T L Peeters
    Abstract:

    Motilin has excitatory effects on the colon of the rabbit and the dog, but little is known of its effect on the human colon. The aim of this study was to investigate the effects induced by motilin and erythromycin A (EMA) on muscle strips and on single cells from primary cultures from human colon. Isotonic contraction was recorded in circular muscle strips from macroscopically normal resection specimens of patients operated on for colonic neoplasm. Agonist-induced intracellular Ca2+ ([Ca2+]i) signalling was studied in primary cultures of colonic smooth-muscle cells using the ratiometric Ca2+ indicator Indo 1, on a laser-scanning confocal epifluorescence microscope. In circular muscle strips, norleucine13-porcine motilin ([Nle13]-pm)and EMA induced tonic contractions with an EC50 of 92 +/- 21 nmol L(-1) and 31 +/- 16 micromol L(-1), respectively. The maximal contraction was 21 +/- 4% (motilin) and 33 +/- 12% (EMA) of the response to 10(-4) mol L(-1) acetylcholine (ACh). The motilin antagonist OHM-11526 (10(-5.5) mol L(-1)) abolished the effects of both [Nle13]-pm and EMA. Neither tetrodotoxin (10(-5.5) mol L(-1)), L-nitro-D-arginine methyl ester (L-NAME) (10(-3.5) mol L(-1)) nor guanethidine (10(-5) mol L(-1)) interfered with the effects of [Nle13]-pm or EMA. [Nle13]-pm (10(-11)-10(-6) mol L(-1)) induced rises of [Ca2+]i in cultured colonic myocytes. At 10(-6) mol L-1, 94% of the cells responded, and half of the cells responded at 1.4 nmol L(-1) [Nle13]-pm. 81% (35/43) and 95% (75/79) responded to EMA (10(-6) mol L(-1)) and acetylcholine (ACh, 10(-4) mol L(-1)), respectively. The motilin antagonist GM-109 inhibited motilin- and EMA-induced [Ca2+]i rises. In the absence of extracellular Ca2+, only 13% (7/52) of the cells responded to [Nle13]-pm (10(-6) mol L(-1)) vs. 90% (47/52) to ACh (10(-4) mol L(-1)). Motilin and EMA have direct excitatory effects on circular smooth muscle from the human colon and these effects are mediated via a smooth-muscle motilin receptor. These findings suggest that motilin may regulate colonic motility and that Motilides may have therapeutic potential for the treatment of colonic hypomotility.

André J. P. M. Smout - One of the best experts on this subject based on the ideXlab platform.

  • effects of a new Motilide abt 229 on gastric emptying and postprandial antroduodenal motility in healthy volunteers
    Alimentary Pharmacology & Therapeutics, 1997
    Co-Authors: M. A. M. T. Verhagen, Melvin Samsom, Bart Maes, Benny Geypens, Yvo Ghoos, André J. P. M. Smout
    Abstract:

    Methods: ABT-229 is a recently developed derivative of erythromycin, devoid of antibiotic activity. We studied the effect of ABT-229 on gastric emptying and postprandial antroduodenal motility in healthy volunteers. Methods: Placebo, 4 and 16 mg ABT-229 were given as a single oral dose to nine healthy volunteers, in a randomized, 3-period crossover design. A solid meal (250 kcal) was given twice, 45 min after drug ingestion and 4 h later. Gastric emptying of each meal was studied using the 13C-octanoic breath test. Antroduodenal motility was recorded during the total 9-h period. Results: After the first meal, both the 4 and 16 mg doses increased the gastric emptying rate to a similar extent. ABT-229 stimulated the contractile motility of the antrum dose-dependently. The half-emptying time and the lag-phase of gastric emptying correlated with the number of pressure waves that were propagated over the antrum and the mean amplitude of antral pressure waves. After the second meal no significant effects of ABT-229 were found. Conclusions: A single dose of the new motilin agonist ABT-229 strongly increases the gastric emptying rate in healthy volunteers by increasing the strength and length of propagation of antral pressure waves. ABT-229 has the potential to become a new prokinetic drug.

  • Effects of a new Motilide, ABT‐229, on gastric emptying and postprandial antroduodenal motility in healthy volunteers
    Alimentary pharmacology & therapeutics, 1997
    Co-Authors: M. A. M. T. Verhagen, Melvin Samsom, Bart Maes, Benny Geypens, Yvo Ghoos, André J. P. M. Smout
    Abstract:

    Methods: ABT-229 is a recently developed derivative of erythromycin, devoid of antibiotic activity. We studied the effect of ABT-229 on gastric emptying and postprandial antroduodenal motility in healthy volunteers. Methods: Placebo, 4 and 16 mg ABT-229 were given as a single oral dose to nine healthy volunteers, in a randomized, 3-period crossover design. A solid meal (250 kcal) was given twice, 45 min after drug ingestion and 4 h later. Gastric emptying of each meal was studied using the 13C-octanoic breath test. Antroduodenal motility was recorded during the total 9-h period. Results: After the first meal, both the 4 and 16 mg doses increased the gastric emptying rate to a similar extent. ABT-229 stimulated the contractile motility of the antrum dose-dependently. The half-emptying time and the lag-phase of gastric emptying correlated with the number of pressure waves that were propagated over the antrum and the mean amplitude of antral pressure waves. After the second meal no significant effects of ABT-229 were found. Conclusions: A single dose of the new motilin agonist ABT-229 strongly increases the gastric emptying rate in healthy volunteers by increasing the strength and length of propagation of antral pressure waves. ABT-229 has the potential to become a new prokinetic drug.