A Potential Treatment of Congenital Sodium Diarrhea in Patients With Activating GUCY2C Mutations
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ABSTRACT: INTRODUCTION:
Gain-of-function mutations in guanylyl cyclase C (GCC) result in persistent diarrhea with perinatal onset. We investigated a specific GCC inhibitor, SSP2518, for its potential to treat this disorder. METHODS:
We investigated the effect of SSP2518 on GCC-mediated intracellular cyclic guanosine monophosphate (cGMP) levels and on GCC-mediated chloride secretion in intestinal organoids from 3 patients with distinct activating GCC mutations and from controls, with and without stimulation of GCC with heat-stable enterotoxin. RESULTS:
Patient-derived organoids had significantly higher basal cGMP levels than control organoids, which were lowered by SSP2518 to levels found in control organoids. In addition, SSP2518 significantly reduced cGMP levels and chloride secretion in patient-derived and control organoids (P < 0.05 for all comparisons) after heat-stable enterotoxin stimulation. DISCUSSION:
We reported in this study that the GCC inhibitor SSP2518 normalizes cGMP levels in intestinal organoids derived from patients with GCC gain-of-function mutations and markedly reduces cystic fibrosis transmembrane conductance regulator–dependent chloride secretion, the driver of persistent diarrhea.
SUBMITTER: van Vugt A
PROVIDER: S-EPMC8604003 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:ObjectiveCongenital sodium diarrhoea (CSD) refers to a form of secretory diarrhoea with intrauterine onset and high faecal losses of sodium without congenital malformations. The molecular basis for CSD remains unknown. We clinically characterised a cohort of infants with CSD and set out to identify disease-causing mutations by genome-wide genetic testing.DesignWe performed whole-exome sequencing and chromosomal microarray analyses in 4 unrelated patients, followed by confirmatory Sanger sequencing of the likely disease-causing mutations in patients and in their family members, followed by functional studies.ResultsWe identified novel de novo missense mutations in GUCY2C, the gene encoding receptor guanylate cyclase C (GC-C) in 4 patients with CSD. One patient developed severe, early-onset IBD and chronic arthritis at 4 years of age. GC-C is an intestinal brush border membrane-bound guanylate cyclase, which functions as receptor for guanylin, uroguanylin and Escherichia coli heat-stable enterotoxin. Mutations in GUCY2C were present in different intracellular domains of GC-C, and were activating mutations that enhanced intracellular cyclic guanosine monophosphate accumulation in a ligand-independent and ligand-stimulated manner, following heterologous expression in HEK293T cells.ConclusionsDominant gain-of-function GUCY2C mutations lead to elevated intracellular cyclic guanosine monophosphate levels and could explain the chronic diarrhoea as a result of decreased intestinal sodium and water absorption and increased chloride secretion. Thus, mutations in GUCY2C indicate a role for this receptor in the pathogenesis of sporadic CSD.
| S-EPMC4975829 | biostudies-literature
Project description:IntroductionIncreased intestinal hydration by activation of the epithelial enzyme linked receptor guanylate cyclase C (GC-C) is a pharmacological principle for treating constipation. Activating mutations in the GUCY2C gene encoding GC-C cause Familial GUCY2C diarrhea syndrome (FGDS) which has been diagnosed with severe dysmotility.AimTo investigate gut motility and hormones before and after a meal in FGDS patients and compare with healthy controls (HC).Subjects and methodsBristol stool chart and stool frequency was assessed. Before and after a meal occlusive and non-occlusive contractions were obtained using ultrasound. A wireless motility capsule (WMC) recorded gut transit time, pH, contractions and pressure. Plasma levels of selected gut hormones were measured at different time points.ResultsThe FGDS patients had 4 (range 1-10) loose stools/day and prolonged total gut transit time compared to HC, 55.5 h vs 28.5 h, respectively,with significantly increased colon transit time. In FGDS patients, pH in duodenum, small bowel and colon was increased and the number of contractions and the intraluminal pressure were significantly decreased, measured by WMC. Ultrasound showed in small bowel increased number of non-occlusive contractions in the FGDS patients. Serotonin (5-HT) plasma levels in the HC peaked 30 min after the meal, while the FGDS patients had no response.ConclusionDespite having diarrhea, the FGDS patients have prolonged transit time through the gut compared to HC, particularly in colon. The reduced number of intestinal contractions and lack of 5-HT release after a meal in FGDS patients surprisingly resemble colonic motility disturbances seen in patients with constipation.
| S-EPMC5619782 | biostudies-literature
Project description:Autosomal-recessive congenital sodium diarrhea (CSD) is characterized by perinatal onset of a persistent watery diarrhea with nonproportionally high fecal sodium excretion. Defective jejunal brush-border Na(+)/H(+) exchange has been reported in three sporadic patients, but the molecular basis of the disease has not been elucidated. We reviewed data from a large cohort of CSD patients (n = 24) and distinguished CSD associated with choanal or anal atresia, hypertelorism, and corneal erosions--i.e., a syndromic form of CSD--occurring in ten families from an isolated form--i.e., classic CSD--presenting in seven families. Patients from both groups have a high risk of mortality due to immediate electrolyte imbalances and complications from long-term parenteral nutrition in the first years of life, but survivors can eventually adapt to partial or complete enteral nutrition. A genome-wide SNP scan was applied and identified a homozygous c.593-1G-->A splicing mutation in SPINT2, encoding a Kunitz-type serine-protease inhibitor, in one extended kindred with syndromic CSD. The same mutation and four distinct, homozygous or compound heterozygous mutations (p.Y163C, c.1A-->T, c.337+2T-->C, c.553+2T-->A) were identified in all syndromic patients. No SPINT2 mutations were found in classic-CSD patients. SPINT2 mutations were associated with loss of protein synthesis or failure to inhibit the serine protease trypsin in vitro. We delineate syndromic CSD as a distinct disease entity caused by SPINT2 loss-of-function mutations. SPINT2 mutations might lead to an excess of yet unknown serine protease activity in affected tissues.
| S-EPMC2668003 | biostudies-literature
Project description:Congenital sodium diarrhea (CSD) refers to an intractable diarrhea of intrauterine onset with high fecal sodium loss. CSD is clinically and genetically heterogeneous. Syndromic CSD is caused by SPINT2 mutations. While we recently described four cases of the non-syndromic form of CSD that were caused by dominant activating mutations in intestinal receptor guanylate cyclase C (GC-C), the genetic cause for the majority of CSD is still unknown. Therefore, we aimed to determine the genetic cause for non-GC-C non-syndromic CSD in 18 patients from 16 unrelated families applying whole-exome sequencing and/or chromosomal microarray analyses and/or direct Sanger sequencing. SLC9A3 missense, splicing and truncation mutations, including an instance of uniparental disomy, and whole-gene deletion were identified in nine patients from eight families with CSD. Two of these nine patients developed inflammatory bowel disease (IBD) at 4 and 16 years of age. SLC9A3 encodes Na(+)/H(+) antiporter 3 (NHE3), which is the major intestinal brush-border Na(+)/H(+) exchanger. All mutations were in the NHE3 N-terminal transport domain, and all missense mutations were in the putative membrane-spanning domains. Identified SLC9A3 missense mutations were functionally characterized in plasma membrane NHE null fibroblasts. SLC9A3 missense mutations compromised NHE3 activity by reducing basal surface expression and/or loss of basal transport function of NHE3 molecules, whereas acute regulation was normal. This study identifies recessive mutations in NHE3, a downstream target of GC-C, as a cause of CSD and implies primary basal NHE3 malfunction as a predisposition for IBD in a subset of patients.
| S-EPMC4634371 | biostudies-literature
Project description:BACKGROUND Familial diarrheas are mostly severe recessive diseases. Here we describe the clinical picture and dominant genetic cause of a novel disease in 32 members of a Norwegian family. The chronic diarrhea is of early onset, relatively mild, and is combined with increased susceptibility to intestinal inflammation, ileus and oesophagitis. METHODS Whole genome SNP-analysis was used to identify a single candidate locus for dominant diarrhea on chromosome 12, followed by sequencing of the GU2CY gene. This encodes guanylyl cyclase 2C, an intestinal receptor for bacterial heat-stable enterotoxins and the peptides uroguanylin and guanylin. Functional studies of a missense mutation were performed after heterologous expression of the mutant receptor in HEK293T cells. The therapeutic response to metformin, an inhibitor of the cystic fibrosis transmembrane regulator (CFTR), was investigated in 5 patients. RESULTS We identified heterozygosity for the first described pathogenic human mutation (c.2519G>T ) in the GUCY2C gene. The mutant receptor showed increased cGMP production in response to its ligands. This may cause hyperactivation of the CFTR and consequent increased chloride and water secretion from the enterocytes, resulting in chronic diarrhea. Treatment with the CFTR-inhibitor metformin significantly reduced the frequency of stools in affected individuals by 34-45%. CONCLUSIONS Increased GC-C signaling disturbs normal bowel function, as shown by the diverse symptoms in our patients, and seems to have a pro-inflammatory effect, either through increased Cl- secretion or additional effects of elevated cellular cGMP. The importance of genetic variants in the GC-C-CFTR-pathway for conditions like Crohn’s disease and IBS should be further explored. Linkage analysis was performed (Allegro v 2.0) using Affymetrix 250K SNP arrays according to the manufacturer's directions on DNA extracted from peripheral blood samples. This accession number contains the data for 25 individuals of family branch A in the article.
2012-04-09 | E-GEOD-31260 | biostudies-arrayexpress
Project description:BACKGROUND Familial diarrheas are mostly severe recessive diseases. Here we describe the clinical picture and dominant genetic cause of a novel disease in 32 members of a Norwegian family. The chronic diarrhea is of early onset, relatively mild, and is combined with increased susceptibility to intestinal inflammation, ileus and oesophagitis. METHODS Whole genome SNP-analysis was used to identify a single candidate locus for dominant diarrhea on chromosome 12, followed by sequencing of the GU2CY gene. This encodes guanylyl cyclase 2C, an intestinal receptor for bacterial heat-stable enterotoxins and the peptides uroguanylin and guanylin. Functional studies of a missense mutation were performed after heterologous expression of the mutant receptor in HEK293T cells. The therapeutic response to metformin, an inhibitor of the cystic fibrosis transmembrane regulator (CFTR), was investigated in 5 patients. RESULTS We identified heterozygosity for the first described pathogenic human mutation (c.2519G>T ) in the GUCY2C gene. The mutant receptor showed increased cGMP production in response to its ligands. This may cause hyperactivation of the CFTR and consequent increased chloride and water secretion from the enterocytes, resulting in chronic diarrhea. Treatment with the CFTR-inhibitor metformin significantly reduced the frequency of stools in affected individuals by 34-45%. CONCLUSIONS Increased GC-C signaling disturbs normal bowel function, as shown by the diverse symptoms in our patients, and seems to have a pro-inflammatory effect, either through increased Cl- secretion or additional effects of elevated cellular cGMP. The importance of genetic variants in the GC-C-CFTR-pathway for conditions like Crohn’s disease and IBS should be further explored.
2012-04-10 | GSE31260 | GEO
Project description:BackgroundCongenital sodium diarrhea (CSD) is a rare enteropathy displaying both broad variability in clinical severity and genetic locus and allelic heterogeneity. Eleven CSD patients were reported so far with SLC9A3 variants that impair the function of the encoded intestinal sodium-proton exchanger 3 (NHE3).MethodsWe report a 4-year-old patient, born prematurely in the 35th week of gestation, with antenatal polyhydramnios and dilated intestinal loops, and with diarrhea of congenital onset, 2-6 times a day, and with polydipsia. She thrived age-appropriately under a normal family diet. Serum sodium levels were repeatedly normal but urinary sodium excretion was low. Exome sequencing revealed compound heterozygous variants in SLC9A3 as the likely cause of the congenital diarrhea.ResultsWhile exome sequencing did not reveal pathogenic or likely pathogenic variants in other genes that cause syndromic or non-syndromic forms of congenital and intractable diarrheas, we identified novel compound heterozygous variants in SLC9A3, a complex allele with two missense changes, NP_004165.2:p.[Ser331Leu;Val449Ile] and in-trans the missense variant p.(Phe451Ser).ConclusionThe clinical phenotype here appears to localize to the milder end of the known CSD spectrum, and the identified variants suggest that this is the twelfth patient reported to date with CSD due to mutations in SLC9A3.
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Project description:Meconium ileus, intestinal obstruction in the newborn, is caused in most cases by CFTR mutations modulated by yet-unidentified modifier genes. We now show that in two unrelated consanguineous Bedouin kindreds, an autosomal-recessive phenotype of meconium ileus that is not associated with cystic fibrosis (CF) is caused by different homozygous mutations in GUCY2C, leading to a dramatic reduction or fully abrogating the enzymatic activity of the encoded guanlyl cyclase 2C. GUCY2C is a transmembrane receptor whose extracellular domain is activated by either the endogenous ligands, guanylin and related peptide uroguanylin, or by an external ligand, Escherichia coli (E. coli) heat-stable enterotoxin STa. GUCY2C is expressed in the human intestine, and the encoded protein activates the CFTR protein through local generation of cGMP. Thus, GUCY2C is a likely candidate modifier of the meconium ileus phenotype in CF. Because GUCY2C heterozygous and homozygous mutant mice are resistant to E. coli STa enterotoxin-induced diarrhea, it is plausible that GUCY2C mutations in the desert-dwelling Bedouin kindred are of selective advantage.
| S-EPMC3376486 | biostudies-literature
Project description:Guanylate cyclase 2C (GC-C), encoded by the GUCY2C gene, is implicated in hereditary early onset chronic diarrhea. Several families with chronic diarrhea symptoms have been identified with autosomal dominant, gain-of-function mutations in GUCY2C. We have identified a Mennonite patient with a novel GUCY2C variant (c.2381A > T; p.Asp794Val) with chronic diarrhea and an extensive maternal family history of chronic diarrhea and bowel dilatation. Functional studies including co-segregation analysis showed that all family members who were heterozygous for this variant had GI-related symptoms. HEK-293 T cells expressing the Asp794Val GC-C variant showed increased cGMP production when stimulated with Escherichia coli heat-stable enterotoxin STp (HST), which was reversed when 5-(3-Bromophenyl)-5,11-dihydro-1,3-dimethyl-1H-indeno[2',1':5,6]pyrido[2,3-d]pyrimidine-2,4,6(3H)-trione (BPIPP; a GC-C inhibitor) was used. In addition, cystic fibrosis transmembrane conductance regulator (CFTR) activity measured with SPQ fluorescence assay was increased in these cells after treatment with HST, indicating a crucial role for CFTR activity in the pathogenesis of this disorder. These results support pathogenicity of the GC-C Asp794Val variant as a cause of chronic diarrhea in this family. Furthermore, this work identifies potential candidate drug, GC-C inhibitor BPIPP, to treat diarrhea caused by this syndrome.
| S-EPMC8251925 | biostudies-literature
Project description:Congenital sodium diarrhea is a rare and life-threatening disorder characterized by a severe, secretory diarrhea containing high concentrations of sodium, leading to hyponatremia and metabolic acidosis. It may occur in isolation or in association with systemic features such as facial dysmorphism, choanal atresia, imperforate anus, and corneal erosions. Mutations in the serine protease inhibitor, Kunitz-Type 2 (SPINT2) gene have been associated with congenital sodium diarrhea and additional syndromic features. We present a child with congenital sodium diarrhea, cleft lip and palate, corneal erosions, optic nerve coloboma, and intermittent exotropia who was found to have biallelic mutations in SPINT2. One mutation, c.488A?>?G, predicting p.(Tyr163Cys), has been previously associated with a syndromic form of congenital sodium diarrhea. The other mutation, c.166_167dupTA, predicting p.(Asn57Thrfs*24) has not previously been reported and is likely a novel pathogenic variant for this disorder. We found only one other report of an optic nerve coloboma associated with SPINT2 mutations and this occurred in a patient with congenital tufting enteropathy. Our patient confirms an association of ocular coloboma with presumed loss of SPINT2 function.
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