Cyst growth in ADPKD is prevented by pharmacological and genetic inhibition of TMEM16A in vivo

Abstract

In autosomal dominant polycystic kidney disease (ADPKD) multiple bilateral renal cysts gradually enlarge, leading to a decline in renal function. Transepithelial chloride secretion through cystic fibrosis transmembrane conductance regulator (CFTR) and TMEM16A (anoctamin 1) are known to drive cyst enlargement. Here we demonstrate that loss of Pkd1 increased expression of TMEM16A and CFTR and Cl − secretion in murine kidneys, with TMEM16A essentially contributing to cyst growth. Upregulated TMEM16A enhanced intracellular Ca 2+ signaling and proliferation of Pkd1-deficient renal epithelial cells. In contrast, increase in Ca 2+ signaling, cell proliferation and CFTR expression was not observed in Pkd1/Tmem16a double knockout mice. Knockout of Tmem16a or inhibition of TMEM16A in vivo by the FDA-approved drugs niclosamide and benzbromarone, as well as the TMEM16A-specific inhibitor Ani9 largely reduced cyst enlargement and abnormal cyst cell proliferation. The present data establish a therapeutic concept for the treatment of ADPKD.

Gender-Dependent Phenotype in Polycystic Kidney Disease Is Determined by Differential Intracellular Ca2+ Signals

“…The KspCreERT2;Pkd1lox;lox system was used to obtain a tamoxifen inducible tubule-specific knockout of Pkd1, as described previously [ 24 , 33 ]. Knockdown of Pkd1 in male and female mice was validated by RT-PCR ( Figure 1 A,B).…”

“…In our previous study we reported a crucial role of the Ca 2+ -activated Cl − channel TMEM16A for the development of polycystic kidneys in Pkd1 −/− mice and in additional in vitro models for ADPKD [ 20 , 23 , 24 ]. We therefore expected to find higher levels of TMEM16A-expression in kidneys from male Pkd1 −/− mice.…”

“…Cell proliferation is enhanced in kidneys from Pkd1 −/− mice [ 24 ]. We compared the cell proliferation in males and females using Ki-67 staining.…”

“…The results show that cell proliferation is enhanced in kidneys of both male and female Pkd1 −/− mice, however, proliferation was more enhanced in males ( Figure 3 A,B). Cellular Ca 2+ levels are intimately related with cell proliferation, and have been shown to be augmented in renal epithelial cells from mice lacking expression of Pkd1, which leads to enhanced expression of TMEM16A [ 23 , 24 , 25 ]. We therefore compared the basal Ca 2+ levels in primary renal epithelial cells from male and female mice.…”

“…Subsequent work identified a contribution of purinergic Ca 2+ signaling to ADPKD [ 17 , 18 , 19 ]. In a series of studies in vitro, in metanephric renal organ cultures, and in mice in vivo, we demonstrated the essential contribution of TMEM16A to renal cyst formation in ADPKD [ 20 , 21 , 22 , 23 , 24 ]. We showed that knockout of Tmem16a or inhibition of TMEM16A in vivo by the FDA-approved drugs such as niclosamide, benzbromarone, and the TMEM16A-specific inhibitor Ani9 largely reduced cyst enlargement and abnormal cyst cell proliferation.…”