Brian J. Murphy

Differential profile of CRF receptor distribution in the rat stomach and duodenum assessed by newly developed CRF receptor antibodies

Peripheral corticotropin‐releasing factor (CRF) receptor ligands inhibit gastric acid secretion and emptying while stimulating gastric mucosal blood flow in rats. Endogenous CRF ligands are expressed in the upper gastrointestinal (GI) tissues pointing to local expression of CRF receptors. We mapped the distribution of CRF receptor type 1 (CRF1) and 2 (CRF2) in the rat upper GI. Polyclonal antisera directed against the C‐terminus of the CRF receptor protein were generated in rabbits and characterized by western blotting and immunofluorescence using CRF1‐ and CRF2‐transfected cell lines and in primary cultured neurons from rat brain cortex. A selective anti‐CRF1 antiserum (4467a‐CRF1) was identified and used in parallel with another antiserum recognizing both CRF1 and CRF2 (4392a‐CRF1&2) to immunostain gastric tissue sections. Antiserum 4467a‐CRF1 demonstrated specific immunostaining in a narrow zone in the upper oxyntic gland within the stomach corpus. Conversely, 4392a‐CRF1&2 labeled cells throughout the oxyntic gland and submucosal blood vessels. Pre‐absorption with the specific antigen peptide blocked immunostaining in all experiments. Doublestaining showed co‐localization of 4392a‐CRF1&2 but not 4467a‐CRF1 immunoreactivity with H/K‐ATPase and somatostatin immunostaining in parietal and endocrine cells of the oxyntic gland. No specific staining was observed in the antrum with either antisera, whereas only antiserum 4392a‐CRF1&2 showed modest immunoreactivity in the duodenal mucosa. Finally, co‐localization of CRF2 and urocortin immunoreactivity was found in the gastric glands. These results indicate that both CRF receptor subtypes are expressed in the rat upper GI tissues with a distinct pattern and regional differences suggesting differential function.

Enhanced Gastrointestinal Motility with Orally Active Ghrelin Receptor Agonists

The orexigenic peptide ghrelin has been shown to have prokinetic activity in the gastrointestinal (GI) system of several species, including humans. In this series of experiments, we have evaluated the prokinetic activity of novel, small-molecule ghrelin receptor (GhrR) agonists after parenteral and peroral dosing in mice and rats. Gastric emptying, small intestinal transport, and fecal output were determined after intraperitoneal and intracerebroventricular dosing of GhrR agonists, using ghrelin as a positive control. These same parameters were evaluated after oral gavage dosing of the synthetic agonists. Regardless of dose route, GhrR agonist treatment increased gastric emptying, small intestinal transit, and fecal output. However, fecal output was only increased by GhrR agonist treatment if mice were able to feed during the stimulatory period. Thus, GhrR agonists can stimulate upper GI motility, and the orexigenic action of the compounds can indirectly contribute to prokinetic activity along the entire GI tract. The orexigenic and prokinetic effects of either ghrelin or small-molecule GhrR agonists were selective for the GhrR because they were absent when evaluated in GhrR knockout mice. We next evaluated the efficacy of the synthetic GhrR agonists dosed in a model of opiate-induced bowel dysfunction induced by a single injection of morphine. Oral dosing of a GhrR agonist normalized GI motility in opiate-induced dysmotility. These data demonstrate the potential utility of GhrR agonists for treating gastrointestinal hypomotility disorders.

Aryl piperazine melanocortin MC4 receptor agonists

Incorporation of substituted phenyl piperazine privileged structures into a known MC4 specific dipeptoid consensus sequence resulted in a series of potent (EC(50)=24 nM) and selective MC4-R agonists. We report the SAR of this series of compounds using in vitro cAMP functional assays in cells transfected with the MC4 or other melancortin receptors.

Body weight regulation by selective MC4 receptor agonists and antagonists.

Abstract: There has been great interest in melanocortin (MC) receptors as targets for the design of novel therapeutics to treat disorders of body weight, such as obesity and cachexia. Both genetic and pharmacological evidence points toward central MC4 receptors as the principal target. Using highly selective peptide tools for the MC4 receptor, which have become available recently, we have provided pharmacological confirmation that central MC4 receptors are the prime mediators of the anorexic and orexigenic effects reported for melanocortin agonists and antagonists, respectively. The current progress with receptor‐selective small molecule agonist and antagonist drugs should enable the therapeutic potential of MC4 receptor activation and inhibition to be assessed in the clinic in the near future.

Central G-Protein Coupled Receptors (GPCR)s as molecular targets for the treatment of obesity: assets, liabilities and development status.

In the last decade, the G-Protein-Coupled Receptor (GPCR) superfamily has emerged as a very promising and enriched source of therapeutic targets for the treatment of obesity. GPCRs represent the largest family of mammalian proteins, with approximately 1000 members. It is estimated that the GPCR family may comprise greater than 1% of the human genome and is the molecular target for approximately 30% of currently marketed drugs. Human GPCRs are modulated by a large variety of ligands, including peptides, lipids, neurotransmitters, nucleotides, ions and external sensory signals such as pheromones, tastes or odors. Many of the above ligands have been implicated in the physiological control of energy balance. This article will examine the biological rationale, assets, identified liabilities and current drug development status of these receptors as anti-obesity drug targets.

Identification of a nonbasic melanin hormone receptor 1 antagonist as an antiobesity clinical candidate.

Identification of MCHR1 antagonists with a preclinical safety profile to support clinical evaluation as antiobesity agents has been a challenge. Our finding that a basic moiety is not required for MCHR1 antagonists to achieve high affinity allowed us to explore structures less prone to off-target activities such as hERG inhibition. We report the SAR evolution of hydroxylated thienopyrimidinone ethers culminating in the identification of 27 (BMS-819881), which entered obesity clinical trials as the phosphate ester prodrug 35 (BMS-830216).