Intestinal GUCY2C prevents TGF-ß secretion coordinating desmoplasia and hyperproliferation in colorectal cancer.

Tumorigenesis is a multistep process that reflects intimate reciprocal interactions between epithelia and underlying stroma. However, tumor-initiating mechanisms coordinating transformation of both epithelial and stromal components are not defined. In humans and mice, initiation of colorectal cancer is universally associated with loss of guanylin and uroguanylin, the endogenous ligands for the tumor suppressor guanylyl cyclase C (GUCY2C), disrupting a network of homeostatic mechanisms along the crypt-surface axis. Here, we reveal that silencing GUCY2C in human colon cancer cells increases Akt-dependent TGF-ß secretion, activating fibroblasts through TGF-ß type I receptors and Smad3 phosphorylation. In turn, activating TGF-ß signaling induces fibroblasts to secrete hepatocyte growth factor (HGF), reciprocally driving colon cancer cell proliferation through cMET-dependent signaling. Elimination of GUCY2C signaling in mice (Gucy2c(-/-)) produces intestinal desmoplasia, with increased reactive myofibroblasts, which is suppressed by anti-TGF-ß antibodies or genetic silencing of Akt. Thus, GUCY2C coordinates intestinal epithelial-mesenchymal homeostasis through reciprocal paracrine circuits mediated by TGF-ß and HGF. In that context, GUCY2C signaling constitutes a direct link between the initiation of colorectal cancer and the induction of its associated desmoplastic stromal niche. The recent regulatory approval of oral GUCY2C ligands to treat chronic gastrointestinal disorders underscores the potential therapeutic opportunity for oral GUCY2C hormone replacement to prevent remodeling of the microenvironment essential for colorectal tumorigenesis.

Linaclotide inhibits colonic nociceptors and relieves abdominal pain via guanylate cyclase-C and extracellular cyclic guanosine 3',5'-monophosphate.

BACKGROUND & AIMS: Linaclotide is a minimally absorbed agonist of guanylate cyclase-C (GUCY2C or GC-C) that reduces symptoms associated with irritable bowel syndrome with constipation (IBS-C). Little is known about the mechanism by which linaclotide reduces abdominal pain in patients with IBS-C. METHODS: We determined the effects of linaclotide on colonic sensory afferents in healthy mice and those with chronic visceral hypersensitivity. We assessed pain transmission by measuring activation of dorsal horn neurons in the spinal cord in response to noxious colorectal distention. Levels of Gucy2c messenger RNA were measured in tissues from mice using quantitative reverse transcription polymerase chain reaction and in situ hybridization. We used human intestinal cell lines to measure release of cyclic guanosine-3',5'-monophosphate (cGMP) by linaclotide. We performed a post-hoc analysis of data from a phase III, double-blind, parallel-group study in which 805 patients with IBS-C were randomly assigned to groups given an oral placebo or 290 µg linaclotide once daily for 26 weeks. We quantified changes in IBS-C symptoms, including abdominal pain. RESULTS: In mice, linaclotide inhibited colonic nociceptors with greater efficacy during chronic visceral hypersensitivity. Intra-colonic administration of linaclotide reduced signaling of noxious colorectal distention to the spinal cord. The colonic mucosa, but not neurons, was found to express linaclotide's target, GC-C. The downstream effector of GC-C, cGMP, was released after administration of linaclotide and also inhibited nociceptors. The effects of linaclotide were lost in Gucy2c(-/-) mice and prevented by inhibiting cGMP transporters or removing the mucosa. During 26 weeks of linaclotide administration, a significantly greater percentage of patients (70%) had at least a 30% reduction in abdominal pain compared with patients given placebo (50%). CONCLUSIONS: We have identified an analgesic mechanism of linaclotide: it activates GC-C expressed on mucosal epithelial cells, resulting in the production and release of cGMP. This extracellular cGMP acts on and inhibits nociceptors, thereby reducing nociception. We also found that linaclotide reduces chronic abdominal pain in patients with IBS-C.

Pharmacology and clinical potential of guanylyl cyclase C agonists in the treatment of ulcerative colitis.

Agonists of the transmembrane intestinal receptor guanylyl cyclase C (GCC) have recently attracted interest as promising human therapeutics. Peptide ligands that can specifically induce GCC signaling in the intestine include endogenous hormones guanylin and uroguanylin, diarrheagenic bacterial enterotoxins (ST), and synthetic drugs linaclotide, plecanatide, and SP-333. These agonists bind to GCC at intestinal epithelial surfaces and activate the receptor's intracellular catalytic domain, an event initiating discrete biological responses upon conversion of guanosine-5'-triphosphate to cyclic guanosine monophosphate. A principal action of GCC agonists in the colon is the promotion of mucosal homeostasis and its dependent barrier function. Herein, GCC agonists are being developed as new medications to treat inflammatory bowel diseases, pathological conditions characterized by mucosal barrier hyperpermeability, abnormal immune reactions, and chronic local inflammation. This review will present important concepts underlying the pharmacology and therapeutic utility of GCC agonists for patients with ulcerative colitis, one of the most prevalent inflammatory bowel disease disorders.

A single gene target of an ETS-family transcription factor determines neuronal CO2-chemosensitivity.

Many animals possess neurons specialized for the detection of carbon dioxide (CO(2)), which acts as a cue to elicit behavioral responses and is also an internally generated product of respiration that regulates animal physiology. In many organisms how such neurons detect CO(2) is poorly understood. We report here a mechanism that endows C. elegans neurons with the ability to detect CO(2). The ETS-5 transcription factor is necessary for the specification of CO(2)-sensing BAG neurons. Expression of a single ETS-5 target gene, gcy-9, which encodes a receptor-type guanylate cyclase, is sufficient to bypass a requirement for ets-5 in CO(2)-detection and transforms neurons into CO(2)-sensing neurons. Because ETS-5 and GCY-9 are members of gene families that are conserved between nematodes and vertebrates, a similar mechanism might act in the specification of CO(2)-sensing neurons in other phyla.

Transmembrane guanylate cyclase in intestinal pathophysiology.

PURPOSE OF REVIEW: Production of cyclic guanosine monophosphate (cGMP) by guanylate cyclase is of critical importance to gastrointestinal physiology. Tight regulation of cGMP concentration is necessary for proper intestinal secretion and intestinal epithelial cell proliferative and apoptotic homeostasis. This review focuses on recent work detailing the role of a subset of transmembrane guanylate cyclases in the pathophysiology of intestinal secretory and motility disorders and intestinal epithelial cell transformation. Also considered is the potential for therapeutic manipulation of intestinal guanylate cyclase/cGMP signaling for the correction of chronic constipation and gastrointestinal cancer. RECENT FINDINGS: Recent work in mice and humans suggests a role for transmembrane guanylate cyclases in intestinal fluid secretion as well as hormonal enteric-renal signaling which mediates postprandial natriuresis. Transmembrane guanylate cyclases are also important in gastrointestinal transit rate and motility. Ongoing clinical trials have found that guanylate cyclase activating peptides are safe and effective in the treatment of constipation-predominant irritable bowel syndrome and chronic constipation. In addition, accumulating evidence indicates that membrane-associated guanylate cyclase receptors regulate intestinal epithelial cell homeostatic proliferation and apoptosis as well as gastrointestinal malignancy. The anticancer activity of cGMP signaling in animal studies suggests additional therapeutic applications for guanylate cyclase agonists. SUMMARY: Progress toward understanding gastrointestinal transmembrane guanylate cyclase/cGMP physiology has recently accelerated due to definitive in-vitro studies and work using gene-targeted animal models and has facilitated the development of safe and effective drugs designed to regulate cGMP production in the intestine. Current work should be directed toward a detailed understanding of cGMP effector pathways and the manner in which subcellular concentrations of cGMP regulate them to influence intestinal health and disease.

Characterization of a novel cell-surface protein expressed on human sperm.

BACKGROUND: Precise sperm-oocyte interaction is a critical event for successful fertilization. However, the identity of molecules involved in this process in humans remains largely unknown. This report describes the identification and characterization of a novel cell-surface protein and its potential role in human sperm-oocyte interaction. METHODS AND RESULTS: We previously identified an orphan guanylyl cyclase receptor (mouse GC-G) highly enriched in mouse testis and involved in sperm activation. By using a comparative genomic approach, we found the homologue gene in human (hGC-G) composed of 21 exons, spanning a minimum of 48 kb on chromosome 10q25. Real-time RT-PCR analysis revealed hGC-G mRNA selectively expressed in testis but with low or no expression in all other tissues examined. Compared with mGC-G, the hGC-G transcript contains three 1-bp deletions and two in-frame termination codons, which results in a short putative receptor-like polypeptide. Western blot analysis with an anti-hGC-G-specific antibody confirmed the protein expression of hGC-G in human sperm lysate. Flow cytometry and confocal immunofluorescence analysis demonstrated the localization of hGC-G protein on the acrosome cap and equatorial segment of mature human sperm. In addition, an integrin-binding Arg-Gly-Asp (RGD) motif was found in the extracellular domain of hGC-G. Pre-incubation of the hGC-G RGD peptide with zona pellucida-free oocytes greatly decreased the binding of human sperm to hamster oocytes, which suggests a role for hGC-G role in sperm-oocyte interaction. CONCLUSIONS: hGC-G is a novel surface protein on human sperm and potentially mediates sperm-oocyte interaction through its RGD-containing motif.

Bacterial heat-stable enterotoxins: translation of pathogenic peptides into novel targeted diagnostics and therapeutics.

Heat-stable toxins (STs) produced by enterotoxigenic bacteria cause endemic and traveler's diarrhea by binding to and activating the intestinal receptor guanylyl cyclase C (GC-C). Advances in understanding the biology of GC-C have extended ST from a diarrheagenic peptide to a novel therapeutic agent. Here, we summarize the physiological and pathophysiological role of GC-C in fluid-electrolyte regulation and intestinal crypt-villus homeostasis, as well as describe translational opportunities offered by STs, reflecting the unique characteristics of GC-C, in treating irritable bowel syndrome and chronic constipation, and in preventing and treating colorectal cancer.

Membrane guanylyl cyclase receptors: an update.

Recent studies have demonstrated key roles for several membrane guanylyl cyclase receptors in the regulation of cell hyperplasia, hypertrophy, migration and extracellular matrix production, all of which having an impact on clinically relevant diseases, including tissue remodeling after injury. Additionally, cell differentiation, and even tumor progression, can be profoundly influenced by one or more of these receptors. Some of these receptors also mediate important communication between the heart and intestine, and the kidney to regulate blood volume and Na+ balance.