Gabriella Coruzzi

Fragment based design of new H4 receptor-ligands with anti-inflammatory properties in vivo

Using a previously reported flexible alignment model we have designed, synthesized, and evaluated a series of compounds at the human histamine H 4 receptor (H 4R) from which 2-(4-methyl-piperazin-1-yl)-quinoxaline ( 3) was identified as a new lead structure for H 4R ligands. Exploration of the structure-activity relationship (SAR) of this scaffold led to the identification of 6,7-dichloro 3-(4-methylpiperazin-1-yl)quinoxalin-2(1 H)-one (VUF 10214, 57) and 2-benzyl-3-(4-methyl-piperazin-1-yl)quinoxaline (VUF 10148, 20) as potent H 4R ligands with nanomolar affinities. In vivo studies in the rat reveal that compound 57 has significant anti-inflammatory properties in the carrageenan-induced paw-edema model.

Gastric antisecretory role and immunohistochemical localization of cannabinoid receptors in the rat stomach

The role of cannabinoid (CB) receptors in the regulation of gastric acid secretion was investigated in the rat by means of functional experiments and by immunohistochemistry. In anaesthetized rats with lumen‐perfused stomach, the non selective CB‐receptor agonist WIN 55,212‐2 (0.30 – 4.00 μmol kg−1, i.v.) and the selective CB1‐receptor agonist HU‐210 (0.03 – 1.50 μmol kg−1, i.v.), dose‐dependently decreased the acid secretion induced by both pentagastrin (30 nmol kg−1 h−1) and 2‐deoxy‐D‐glucose (1.25 mmol kg−1, i.v.). By contrast, neither WIN 55,212‐2 (1 – 4 μmol kg−1, i.v.) nor HU‐210 (0.03 – 1.50 μmol kg−1, i.v.) did modify histamine‐induced acid secretion (20 μmol kg−1 h−1). The selective CB2‐receptor agonist JWH‐015 (3 – 10 μmol kg−1, i.v.) was ineffective. The gastric antisecretory effects of WIN 55,212‐2 and HU‐210 on pentagastrin‐induced acid secretion were prevented by the selective CB1‐receptor antagonist SR141716A (0.65 μmol kg−1, i.v.) and unaffected by the selective CB2‐receptor antagonist SR144528 (0.65 – 2 μmol kg−1, i.v.). Bilateral cervical vagotomy and ganglionic blockade with hexamethonium (10 mg kg−1, i.v., followed by continuous infusion of 10 mg kg−1 h−1) significantly reduced, but not abolished, the maximal inhibitory effect of HU‐210 (0.3 μmol kg−1, i.v.) on pentagastrin‐induced acid secretion; by contrast, pretreatment with atropine (1 mg kg−1, i.v.) did not modify the antisecretory effect of HU‐210. Immunoreactivity to the CB1 receptor was co‐localized with that of the cholinergic marker choline acetyltransferase in neural elements innervating smooth muscle, mucosa and submucosal blood vessels of rat stomach fundus, corpus and antrum. In contrast, CB2 receptor‐like immunoreactivity was not observed. These results indicate that gastric antisecretory effects of cannabinoids in the rat are mediated by suppression of vagal drive to the stomach through activation of CB1 receptors, located on pre‐ and postganglionic cholinergic pathways. However, the ineffectiveness of atropine in reducing the effect of HU‐210 suggests that the release of non cholinergic excitatory neurotransmitters may be regulated by CB1 receptors.

Discovery of Quinazolines as Histamine H4 Receptor Inverse Agonists Using a Scaffold Hopping Approach

From a series of small fragments that was designed to probe the histamine H(4) receptor (H(4)R), we previously described quinoxaline-containing fragments that were grown into high affinity H(4)R ligands in a process that was guided by pharmacophore modeling. With a scaffold hopping exercise and using the same in silico models, we now report the identification and optimization of a series of quinazoline-containing H(4)R compounds. This approach led to the discovery of 6-chloro-N-(furan-3-ylmethyl)2-(4-methylpiperazin-1-yl)quinazolin-4-amine (VUF10499, 54) and 6-chloro-2-(4-methylpiperazin-1-yl)-N-(thiophen-2-ylmethyl)quinazolin-4-amine (VUF10497, 55) as potent human H(4)R inverse agonists (pK(i) = 8.12 and 7.57, respectively). Interestingly, both compounds also possess considerable affinity for the human histamine H(1) receptor (H(1)R) and therefore represent a novel class of dual action H(1)R/H(4)R ligands, a profile that potentially leads to added therapeutic benefit. Compounds from this novel series of quinazolines are antagonists at the rat H(4)R and were found to possess anti-inflammatory properties in vivo in the rat.

Cholinergic-like effects of the new histamine H2-receptor antagonist ranitidine

The new H2-receptor blocker ranitidine, together with the effect on histamine H2-receptors, possesses a series of cholinergic-like actions: it provokes atropine-sensitive contractions of several isolated smooth muscle preparations from different animal species and it potentiates the stimulant effect of acetylcholine. Moreover it contracts human lower esophageal sphincterin vivo, an effect which is completely prevented by small doses of atropine. Finally, ranitidine potentiates the stimulant effect of bethanechol and of carbachol on salivary glands of the rat while leaving unaffected the secretagogue effect of physalaemin which is known to be completely independent of the cholinergic system. In thein vivo experiments the doses of ranitidine capable of eliciting cholinergic-like effects were of the same order of magnitude as those necessary to cause the H2-receptor blockade.

Morphological and functional alterations of the myenteric plexus in rats with TNBS-induced colitis

The 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced model of experimental colitis was used to investigate the time-course of alterations in enteric neurotransmission and/or smooth muscle function that occur in chronic inflammation. Myenteric plexus morphology (immunocytochemical markers), functional integrity of cholinergic neurons (3H-choline uptake, acetylcholine release and contractile response to electrical field stimulation) and smooth muscle integrity (contractile response to exogenous acetylcholine) were determined 2, 7, 15, and 30 days after TNBS treatment. In TNBS-treated rats extensive ulcerations of the mucosa and/or the submucosa and increase in colonic weights were accompanied by significant reduction in 3H-choline uptake, acetylcholine release and contractile response to stimulation of enteric nerves. These changes were maximal 7 and 15 days after TNBS treatment. Immunocytochemical marker (PGP 9.5, SNAP 25, synaptophysin and S100 protein) expression was absent in necrotic areas of colons removed 7 days post-injury and partially reduced in colons removed 15 days after TNBS treatment. By contrast, the contractile response to exogenous acetylcholine was significantly increased after 7 days in both inflamed and uninflamed regions and returned to control values by day 30. Likewise, an almost complete recovery of neural cholinergic function and of myenteric plexus morphology was observed 30 days after TNBS treatment. These data suggest that TNBS-induced colitis is associated with progressive and selective alterations in myenteric plexus structure and function, with consequent reduction of cholinergic neurotransmission and abnormality in colonic contractility. The reversibility of myenteric plexus disruption is a clear indication of neuronal plasticity within enteric nervous system as an adaptative mechanism against inflammatory challenges.

Synthesis and QSAR of quinazoline sulfonamides as highly potent human histamine H4 receptor inverse agonists.

Hit optimization of the class of quinazoline containing histamine H(4) receptor (H(4)R) ligands resulted in a sulfonamide substituted analogue with high affinity for the H(4)R. This moiety leads to improved physicochemical properties and is believed to probe a distinct H(4)R binding pocket that was previously identified using pharmacophore modeling. By introducing a variety of sulfonamide substituents, the H(4)R affinity was optimized. The interaction of the new ligands, in combination with a set of previously published quinazoline compounds, was described by a QSAR equation. Pharmacological studies revealed that the sulfonamide analogues have excellent H(4)R affinity and behave as inverse agonists at the human H(4)R. In vivo evaluation of the potent 2-(6-chloro-2-(4-methylpiperazin-1-yl)quinazoline-4-amino)-N-phenylethanesulfonamide (54) (pK(i) = 8.31 +/- 0.10) revealed it to have anti-inflammatory activity in an animal model of acute inflammation.

An update on histamine H3 receptors and gastrointestinal functions

The distribution and functions of histamine H3 receptors in the gastrointestinal tract is reviewed with particular reference to the effects on gastric acid secretion, mucosal protection, and intestinal motility. Histamine H3 receptor activation has negative effects on acid secretion induced by indirect secretagogues in cats, dogs, and rabbits; less clear effects were found in rats. An inhibitory effect on histamine release induced by different stimuli was observed in rats, rabbits, and dogs after H3 receptor agonists, thus supporting the idea that H3 receptors occur in ECL cells. (R)-α-methylhistamine has a marked protective effect against gastric lesions induced by ethanol in rats, being slightly less effective against aspirin and stress. H3 receptor activation decreases the intestinal motility induced by electrical stimulation in a variety of gut preparations, reducing both cholinergic and NANC neurotransmitter release. In this tissue the inhibitory effects mediated by histamine H3 receptors seem to be coupled, via a G protein, to a restriction of Ca2+ access into the nerve terminal; other mechanisms, however, have been suggested in the gastric mucosa. Histamine H3 receptors have already been subdivided into two receptor subtypes, H3A and H3B, the former being the subtype predominant in the gastrointestinal tissue. The increasing availability of selective agonists and antagonists of H3 receptors will unravel possible novel actions and physiological roles of histamine.

The Histamine H3 Receptor

Histamine is widely distributed in the body, although with marked quantitative differences in the various species and tissues, and it produces a variety of biological effects by interacting with specific receptors on the surface of target cells.

Pharmacological characterization of the new histamine H4 receptor agonist VUF 8430

Background and purpose:  We compare the pharmacological profiles of a new histamine H4 receptor agonist 2‐(2‐guanidinoethyl)isothiourea (VUF 8430) with that of a previously described H4 receptor agonist, 4‐methylhistamine.

Histamine H3 receptors regulate acetylcholine release from the guinea pig ileum myenteric plexus

The effect of selective histamine H3-receptor agonists and antagonists on the acetylcholine release from peripheral nerves was evaluated in the guinea pig longitudinal muscle-myenteric plexus preparations, preloaded with (3H)choline. In the presence of H1 and H2 blockade, histamine (10(-7)-10(-4) M) and (R)-alpha-methylhistamine (10(-8)-10(-6) M) inhibited the electrically-evoked acetylcholine release, being (R)-alpha-methylhistamine more active than histamine, but behaving as a partial agonist. The effect of histamine was completely reversed by selective H3-blocking drugs, thioperamide and impromidine, while only submaximal doses of (R)-alpha-methylhistamine were antagonized. Furthermore, thioperamide and impromidine enhanced the electrically-evoked acetylcholine release. On the contrary, the new H3-blocker, HST-7, was found substantially ineffective, both as histamine antagonist and as acetylcholine overflow enhancer. These data suggest that histamine exerts an inhibitory control on the acetylcholine release from intestinal cholinergic nerves through the activation of H3 receptors.