Guanylate cyclase 2C agonism corrects CFTR mutants
Kavisha Arora, Yunjie Huang, Kyushik Mun, Sunitha Yarlagadda, Nambirajan Sundaram, Marco M. Kessler, Gerhard Hannig, Caroline B. Kurtz, Inmaculada Silos-Santiago, Michael Helmrath, Joseph J. Palermo, John P. Clancy, Kris A. Steinbrecher, Anjaparavanda P. Naren
Kavisha Arora, Yunjie Huang, Kyushik Mun, Sunitha Yarlagadda, Nambirajan Sundaram, Marco M. Kessler, Gerhard Hannig, Caroline B. Kurtz, Inmaculada Silos-Santiago, Michael Helmrath, Joseph J. Palermo, John P. Clancy, Kris A. Steinbrecher, Anjaparavanda P. Naren
View: Text | PDF
Research Article Gastroenterology

Guanylate cyclase 2C agonism corrects CFTR mutants

Abstract

Cystic fibrosis (CF) is a genetic disorder in which epithelium-generated fluid flow from the lung, intestine, and pancreas is impaired due to mutations disrupting CF transmembrane conductance regulator (CFTR) channel function. CF manifestations of the pancreas and lung are present in the vast majority of CF patients, and 15% of CF infants are born with obstructed gut or meconium ileus. However, constipation is a significantly underreported outcome of CF disease, affecting 47% of the CF patients, and management becomes critical in the wake of increasing life span of CF patients. In this study, we unraveled a potentially novel molecular role of a membrane-bound cyclic guanosine monophosphate–synthesizing (cGMP-synthesizing) intestinal enzyme, guanylate cyclase 2C (GCC) that could be targeted to ameliorate CF-associated intestinal fluid deficit. We demonstrated that GCC agonism results in functional rescue of murine F508del/F508del and R117H/R117H Cftr and CFTR mutants in CF patient–derived intestinal spheres. GCC coexpression and activation facilitated processing and ER exit of F508del CFTR and presented a potentially novel rescue modality in the intestine, similar to the CF corrector VX-809. Our findings identify GCC as a biological CFTR corrector and potentiator in the intestine.

Authors

Kavisha Arora, Yunjie Huang, Kyushik Mun, Sunitha Yarlagadda, Nambirajan Sundaram, Marco M. Kessler, Gerhard Hannig, Caroline B. Kurtz, Inmaculada Silos-Santiago, Michael Helmrath, Joseph J. Palermo, John P. Clancy, Kris A. Steinbrecher, Anjaparavanda P. Naren

×

Figure 3

Long-term exposure to STc corrects and rescues F508del CFTR function in enterospheres.

Options: View larger image (or click on image) Download as PowerPoint
Long-term exposure to STc corrects and rescues F508del CFTR function in ...
(A) Representative images of F508del/F508del Cftr enterospheres depict secretion under various treatment conditions: (top row) STc (0, 50, 500, and 1,000 nM, 24 h), (middle) STc (0, 50, 500, and 1,000 nM, 24 h) + CFTRinh-172 (20 μM, 30 min), and (bottom) STc (0, 50, 500, and 1,000 nM, 24 h) + VX-809 (2 μM, 24 h). (B) Dot plot represents quantitation of fluid secretion in enterospheres corresponding to conditions from A. Data ± SEM was calculated from n = 14–40 organoids per condition. This experiment was repeated in a total of 5 mice. Lower left panel depicts fluid secretion in WT/WT Cftr enterospheres and quantitated and represented as a dot plot graph. P-value by ANOVA with Bonferroni’s multiple comparisons test. (C) Confocal images of F508del/F508del Cftr intestinal spheres treated and nontreated with STc (50 nM, 24 h) showing CFTR-specific immunostaining (green) and fluorescent labeling corresponding to F-actin (red) and nuclear counterstain (blue). (D) Representative images of R117H/R117H Cftr enterospheres depict secretion in response to forskolin stimulation under various treatment conditions: STc (50 nM, 24 h), VX-809 (2 μM, 24 h), VX-770 (2 μM, 24 h), STc + VX-809, and STc + VX-770. (E) Bar graph depicts fluid secretion calculated from the above-described treatment conditions. All data are represented by mean ± SEM, calculated from n = 25–32 organoids per condition repeated in 2 mice P value by ANOVA with Bonferroni’s multiple comparisons test.