Guanylate cyclase-C/cGMP: an emerging pathway in the regulation of visceral pain

Activation of guanylate cyclase-C (GC-C) expressed predominantly on intestinal epithelial cells by guanylin, uroguanylin or the closely related GC-C agonist peptide, linaclotide, stimulates generation and release of cyclic guanosine-3’,5’-monophosphate (cGMP). Evidence that the visceral analgesic effects of linaclotide are mediated by a novel, GC-C-dependent peripheral sensory mechanism was first demonstrated in animal models of visceral pain. Subsequent studies with uroguanylin or linaclotide have confirmed the activation of a GC-C/cGMP pathway leading to increased submucosal cGMP mediated by cGMP efflux pumps, which modulates intestinal nociceptor function resulting in peripheral analgesia. These effects can be reproduced by the addition of exogenous cGMP and support a role for GC-C/cGMP signaling in the regulation of visceral sensation, a physiological function that has not previously been linked to the GC-C/cGMP pathway. Notably, targeting the GC-C/cGMP pathway for treatment of gastrointestinal pain and abdominal sensory symptoms has now been validated in the clinic. In 2012, linaclotide was approved in the United States and European Union for the treatment of adult patients with irritable bowel syndrome with constipation

Genetically engineered E. coli Nissle attenuates hyperammonemia and prevents memory impairment in bile‐duct ligated rats

Hyperammonemia associated with chronic liver disease (CLD) is implicated in the pathogenesis of hepatic encephalopathy (HE). The gut is a major source of ammonia production that contributes to hyperammonemia in CLD and HE and remains the primary therapeutic target for lowering hyperammonemia. As an ammonia‐lowering strategy, Escherichia coli Nissle 1917 bacterium was genetically modified to consume and convert ammonia to arginine (S‐ARG). S‐ARG was further modified to additionally synthesize butyrate (S‐ARG+BUT). Both strains were evaluated in bile‐duct ligated (BDL) rats; experimental model of CLD and HE. Methods One‐week post‐surgery, BDLs received non‐modified EcN (EcN), S‐ARG, S‐ARG+BUT (3x1011 CFU/day) or vehicle until sacrifice at 3‐ or 5‐weeks. Plasma (ammonia/pro‐inflammatory/liver‐function), liver fibrosis (hydroxyproline), liver mRNA (pro‐inflammatory/fibrogenic/anti‐apoptotic) and colon mRNA (pro‐inflammatory) biomarkers were measured post‐sacrifice. Memory, motor‐coordination, muscle‐strength, and locomotion were assessed at 5‐weeks. Results In BDL‐Veh rats, hyperammonemia developed at 3‐ and further increased at 5‐weeks. This rise was prevented by S‐ARG and S‐ARG+BUT, whereas EcN was ineffective. Memory impairment was prevented only in S‐ARG+BUT vs BDL‐Veh. Systemic inflammation (IL‐10/MCP‐1/endotoxin) increased at 3‐ and 5‐weeks in BDL‐Veh. S‐ARG+BUT attenuated inflammation at both timepoints (except 5‐week endotoxin) vs BDL‐Veh, whereas S‐ARG only attenuated IP‐10 and MCP‐1 at 3‐weeks. Circulating (ALT/AST/ALP/GGT/albumin/bilirubin) and gene expression liver‐function markers (IL‐10/IL‐6/IL‐1β/TGF‐β/α‐SMA/collagen‐1α1/Bcl‐2) were not normalized by either strain. Colonic mRNA (TNF‐α/IL‐1β/occludin) markers were attenuated by synthetic strains at both timepoints vs BDL‐Veh. Conclusion S‐ARG and S‐ARG+BUT attenuated hyperammonemia, with S‐ARG+BUT additional memory protection likely due to greater anti‐inflammatory effect. These innovative strategies, particularly S‐ARG+BUT, have potential to prevent HE

Demonstration of differences in colonic volumes, transit, chyme consistency and response to psyllium between healthy and constipated subjects using magnetic resonance imaging

Background: In functional gastrointestinal disorders a lack of objective biomarkers limits evaluation of underlying mechanisms. We aimed to demonstrate the utility of Magnetic Resonance Imaging (MRI) for this task using psyllium, an effective constipation treatment, in patients and controls. Methods: Two crossover studies: 1) adults without constipation (controls, n=9) took three treatments in randomised order for 6 days - maltodextrin (placebo), psyllium 3.5g t.d.s and 7g t.d.s.; 2) adults with chronic constipation (patients, n=20) took placebo and psyllium 7g t.d.s. for 6 days. MRI was performed fasting and postprandially on day 6. Measurements included small bowel and ascending colon water content, colonic volume, transit time and MR relaxometry (T1, T2) to assess colonic chyme. Stool water percentage was measured. Results: 7g psyllium t.d.s. increased fasting colonic volumes in controls from median 372mL (IQR 284-601) to 578 mL (IQR 510-882), and in patients from median 831mL (IQR 745–934) to 1104mL (847–1316),

Demonstration of differences in colonic volumes, transit, chyme consistency and response to psyllium between healthy and constipated subjects using magnetic resonance imaging

Background: In functional gastrointestinal disorders a lack of objective biomarkers limits evaluation of underlying mechanisms. We aimed to demonstrate the utility of Magnetic Resonance Imaging (MRI) for this task using psyllium, an effective constipation treatment, in patients and controls. Methods: Two crossover studies: 1) adults without constipation (controls, n=9) took three treatments in randomised order for 6 days - maltodextrin (placebo), psyllium 3.5g t.d.s and 7g t.d.s.; 2) adults with chronic constipation (patients, n=20) took placebo and psyllium 7g t.d.s. for 6 days. MRI was performed fasting and postprandially on day 6. Measurements included small bowel and ascending colon water content, colonic volume, transit time and MR relaxometry (T1, T2) to assess colonic chyme. Stool water percentage was measured. Results: 7g psyllium t.d.s. increased fasting colonic volumes in controls from median 372mL (IQR 284-601) to 578 mL (IQR 510-882), and in patients from median 831mL (IQR 745–934) to 1104mL (847–1316), P<0.05). Mean postprandial small bowel water was higher in controls and patients after 7g psyllium t.d.s. vs. placebo. Whole gut transit was slower in patients than controls (P <0.05). T1 of the descending colon chyme (fasting) was lower in patients [213ms, 176–420] than controls [440ms, 352–884, P <0.05] on placebo, but increased by 7 g psyllium t.d.s. [590ms, 446–1338), P<0.001]. Descending colon T1 correlated with baseline stool water content and stool frequency on treatment. Conclusions and Inferences: MRI measurements can objectively demonstrate the mode of action of therapy targeting intestinal fluid content in constipation

A 12-Week, Randomized, Controlled Trial with a 4-Week Randomized Withdrawal Period to Evaluate the Efficacy and Safety of Linaclotide in Irritable Bowel Syndrome with Constipation

Objectives: Linaclotide is a minimally absorbed guanylate cyclase-C agonist. The objective of this trial was to determine the efficacy and safety of linaclotide in patients with irritable bowel syndrome with constipation (IBS-C). Methods: This phase 3, double-blind, parallel-group, placebo-controlled trial randomized IBS-C patients to placebo or 290 μg oral linaclotide once daily in a 12-week treatment period, followed by a 4-week randomized withdrawal (RW) period. There were four primary end points, the Food and Drug Administration's (FDA's) primary end point for IBS-C (responder: improvement of ≥30% in average daily worst abdominal pain score and increase by ≥1 complete spontaneous bowel movement (CSBM) from baseline (same week) for at least 50% of weeks assessed) and three other primary end points, based on improvements in abdominal pain and CSBMs for 9/12 weeks. Adverse events (AEs) were monitored. Results: The trial evaluated 800 patients (mean age=43.5 years, female=90.5%, white=76.9%). The FDA end point was met by 136/405 linaclotide-treated patients (33.6%), compared with 83/395 placebo-treated patients (21.0%) (P<0.0001) (number needed to treat: 8.0, 95% confidence interval: 5.4, 15.5). A greater percentage of linaclotide patients, compared with placebo patients, reported for at least 6/12 treatment period weeks, a reduction of ≥30% in abdominal pain (50.1 vs. 37.5%, P=0.0003) and an increase of ≥1 CSBM from baseline (48.6 vs. 29.6%, P<0.0001). A greater percentage of linaclotide patients vs. placebo patients were also responders for the other three primary end points (P<0.05). Significantly greater improvements were seen in linaclotide vs. placebo patients for all secondary end points (P<0.001). During the RW period, patients remaining on linaclotide showed sustained improvement; patients re-randomized from linaclotide to placebo showed return of symptoms, but without worsening of symptoms relative to baseline. Diarrhea, the most common AE, resulted in discontinuation of 5.7% of linaclotide and 0.3% of placebo patients. Conclusions: Linaclotide significantly improved abdominal pain and bowel symptoms associated with IBS-C for at least 12 weeks; there was no worsening of symptoms compared with baseline following cessation of linaclotide during the RW period

Detectable clonal mosaicism and its relationship to aging and cancer

In an analysis of 31,717 cancer cases and 26,136 cancer-free controls from 13 genome-wide association studies, we observed large chromosomal abnormalities in a subset of clones in DNA obtained from blood or buccal samples. We observed mosaic abnormalities, either aneuploidy or copy-neutral loss of heterozygosity, of >2 Mb in size in autosomes of 517 individuals (0.89%), with abnormal cell proportions of between 7% and 95%. In cancer-free individuals, frequency increased with age, from 0.23% under 50 years to 1.91% between 75 and 79 years (P = 4.8 × 10(-8)). Mosaic abnormalities were more frequent in individuals with solid tumors (0.97% versus 0.74% in cancer-free individuals; odds ratio (OR) = 1.25; P = 0.016), with stronger association with cases who had DNA collected before diagnosis or treatment (OR = 1.45; P = 0.0005). Detectable mosaicism was also more common in individuals for whom DNA was collected at least 1 year before diagnosis with leukemia compared to cancer-free individuals (OR = 35.4; P = 3.8 × 10(-11)). These findings underscore the time-dependent nature of somatic events in the etiology of cancer and potentially other late-onset diseases

Female chromosome X mosaicism is age-related and preferentially affects the inactivated X chromosome

To investigate large structural clonal mosaicism of chromosome X, we analysed the SNP microarray intensity data of 38,303 women from cancer genome-wide association studies (20,878 cases and 17,425 controls) and detected 124 mosaic X events42Mb in 97 (0.25%) women. Here we show rates for X-chromosome mosaicism are four times higher than mean autosomal rates; X mosaic events more often include the entire chromosome and participants with X events more likely harbour autosomal mosaic events. X mosaicism frequency increases with age (0.11% in 50-year olds; 0.45% in 75-year olds), as reported for Y and autosomes. Methylation array analyses of 33 women with X mosaicism indicate events preferentially involve the inactive X chromosome. Our results provide further evidence that the sex chromosomes undergo mosaic events more frequently than autosomes, which could have implications for understanding the underlying mechanisms of mosaic events and their possible contribution to risk for chronic diseases

Female chromosome X mosaicism is age-related and preferentially affects the inactivated X chromosome

To investigate large structural clonal mosaicism of chromosome X, we analysed the SNP microarray intensity data of 38,303 women from cancer genome-wide association studies (20,878 cases and 17,425 controls) and detected 124 mosaic X events >2 Mb in 97 (0.25%) women. Here we show rates for X-chromosome mosaicism are four times higher than mean autosomal rates; X mosaic events more often include the entire chromosome and participants with X events more likely harbour autosomal mosaic events. X mosaicism frequency increases with age (0.11% in 50-year olds; 0.45% in 75-year olds), as reported for Y and autosomes. Methylation array analyses of 33 women with X mosaicism indicate events preferentially involve the inactive X chromosome. Our results provide further evidence that the sex chromosomes undergo mosaic events more frequently than autosomes, which could have implications for understanding the underlying mechanisms of mosaic events and their possible contribution to risk for chronic diseases