scholarly journals High-Throughput Phenotypic Screening of Kinase Inhibitors to Identify Drug Targets for Polycystic Kidney Disease

2017 ◽  
Vol 22 (8) ◽  
pp. 974-984 ◽  
Author(s):  
Tijmen H. Booij ◽  
Hester Bange ◽  
Wouter N. Leonhard ◽  
Kuan Yan ◽  
Michiel Fokkelman ◽  
...  
Keyword(s):  
Kidney Disease ◽  
Signaling Pathways ◽  
Polycystic Kidney Disease ◽  
Kinase Inhibitors ◽  
Renal Cysts ◽  
In Vitro Assays ◽  
Phenotypic Change ◽  
Polycystic Kidney ◽  
Culture Model

Polycystic kidney disease (PKD) is a prevalent disorder characterized by renal cysts that lead to kidney failure. Various signaling pathways have been targeted to stop disease progression, but most interventions still focus on alleviating PKD-associated symptoms. The mechanistic complexity of the disease, as well as the lack of functional in vitro assays for compound testing, has made drug discovery for PKD challenging. To identify modulators of PKD, Pkd1–/– kidney tubule epithelial cells were applied to a scalable and automated 3D cyst culture model for compound screening, followed by phenotypic profiling to determine compound efficacy. We used this screening platform to screen a library of 273 kinase inhibitors to probe various signaling pathways involved in cyst growth. We show that inhibition of several targets, including aurora kinase, CDK, Chk, IGF-1R, Syk, and mTOR, but, surprisingly, not PI3K, prevented forskolin-induced cyst swelling. Additionally, we show that multiparametric phenotypic classification discriminated potentially undesirable (i.e., cytotoxic) compounds from molecules inducing the desired phenotypic change, greatly facilitating hit selection and validation. Our findings show that a pathophysiologically relevant 3D cyst culture model of PKD coupled to phenotypic profiling can be used to identify potentially therapeutic compounds and predict and validate molecular targets for PKD.

scholarly journals Ouabain activates the Na-K-ATPase signalosome to induce autosomal dominant polycystic kidney disease cell proliferation

2011 ◽  
Vol 301 (4) ◽  
pp. F897-F906 ◽  
Author(s):  
Anh-Nguyet T. Nguyen ◽  
Kyle Jansson ◽  
Gladis Sánchez ◽  
Madhulika Sharma ◽  
Gail A. Reif ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Kidney Disease ◽  
Epithelial Cells ◽  
Polycystic Kidney Disease ◽  
Kinase Inhibitors ◽  
Autosomal Dominant ◽  
Renal Cysts ◽  
Cell Nuclei ◽  
Polycystic Kidney ◽  
Autosomal Dominant Polycystic Kidney

The Na-K-ATPase is part of a cell signaling complex, the Na-K-ATPase signalosome, which upon activation by the hormone ouabain regulates the function of different cell types. We previously showed that ouabain induces proliferation of epithelial cells derived from renal cysts of patients with autosomal dominant polycystic kidney disease (ADPKD cells). Here, we investigated the signaling pathways responsible for mediating the effects of ouabain in these cells. Incubation of ADPKD cells with ouabain, in concentrations similar to those found in blood, stimulated phosphorylation of the epidermal growth factor receptor (EGFR) and promoted its association to the Na-K-ATPase. In addition, ouabain activated the kinase Src, but not the related kinase Fyn. Tyrphostin AG1478 and PP2, inhibitors of EGFR and Src, respectively, blocked ouabain-dependent ADPKD cell proliferation. Treatment of ADPKD cells with ouabain also caused phosphorylation of the caveolar protein caveolin-1, and disruption of cell caveolae with methyl-β-cyclodextrin prevented Na-K-ATPase-EGFR interaction and ouabain-induced proliferation of the cells. Downstream effects of ouabain in ADPKD cells included activation of B-Raf and MEK and phosphorylation of the extracellular regulated kinase ERK, which translocated into the ADPKD cell nuclei. Finally, ouabain reduced expression of the cyclin-dependent kinase inhibitors p21 and p27, which are suppressors of cell proliferation. Different from ADPKD cells, ouabain showed no significant effect on B-Raf, p21, and p27 in normal human kidney epithelial cells. Altogether, these results identify intracellular pathways of ouabain-dependent Na-K-ATPase-mediated signaling in ADPKD cells, including EGFR-Src-B-Raf-MEK/ERK, and establish novel mechanisms involved in ADPKD cell proliferation.

scholarly journals In vitro 3D phenotypic drug screen identifies celastrol as an effective in vivo inhibitor of polycystic kidney disease

2019 ◽  
Vol 12 (8) ◽  
pp. 644-653 ◽  
Author(s):  
Tijmen H Booij ◽  
Wouter N Leonhard ◽  
Hester Bange ◽  
Kuan Yan ◽  
Michiel Fokkelman ◽  
...  
Keyword(s):  
Kidney Disease ◽  
Polycystic Kidney Disease ◽  
Genetic Disorder ◽  
Screening Method ◽  
Renal Cysts ◽  
Cyst Formation ◽  
Polycystic Kidney ◽  
Cyst Growth

Abstract Polycystic kidney disease (PKD) is a prevalent genetic disorder, characterized by the formation of kidney cysts that progressively lead to kidney failure. The currently available drug tolvaptan is not well tolerated by all patients and there remains a strong need for alternative treatments. The signaling rewiring in PKD that drives cyst formation is highly complex and not fully understood. As a consequence, the effects of drugs are sometimes difficult to predict. We previously established a high throughput microscopy phenotypic screening method for quantitative assessment of renal cyst growth. Here, we applied this 3D cyst growth phenotypic assay and screened 2320 small drug-like molecules, including approved drugs. We identified 81 active molecules that inhibit cyst growth. Multi-parametric phenotypic profiling of the effects on 3D cultured cysts discriminated molecules that showed preferred pharmacological effects above genuine toxicological properties. Celastrol, a triterpenoid from Tripterygium Wilfordii, was identified as a potent inhibitor of cyst growth in vitro. In an in vivo iKspCre-Pkd1lox,lox mouse model for PKD, celastrol inhibited the growth of renal cysts and maintained kidney function.

scholarly journals Applications of Herbal Medicine to Treat Autosomal Dominant Polycystic Kidney Disease

2021 ◽  
Vol 12 ◽  
Author(s):  
Guangying Shao ◽  
Shuai Zhu ◽  
Baoxue Yang
Keyword(s):  
Kidney Disease ◽  
Polycystic Kidney Disease ◽  
Autosomal Dominant ◽  
Herbal Medicines ◽  
Renal Cysts ◽  
Cordyceps Sinensis ◽  
Polycystic Kidney ◽  
Autosomal Dominant Polycystic Kidney

Autosomal dominant polycystic kidney disease (ADPKD) is a common hereditary kidney disease, which is featured by progressively enlarged bilateral fluid-filled cysts. Enlarging cysts destroy the structure of nephrons, ultimately resulting in the loss of renal function. Eventually, ADPKD develops into end-stage renal disease (ESRD). Currently, there is no effective drug therapy that can be safely used clinically. Patients progressed into ESRD usually require hemodialysis and kidney transplant, which is a heavy burden on both patients and society. Therefore, looking for effective therapeutic drugs is important for treating ADPKD. In previous studies, herbal medicines showed their great effects in multiple diseases, such as cancer, diabetes and mental disorders, which also might play a role in ADPKD treatment. Currently, several studies have reported that the compounds from herbal medicines, such as triptolide, curcumin, ginkolide B, steviol, G. lucidum triterpenoids, Celastrol, saikosaponin-d, Sparganum stoloniferum Buch.-Ham and Cordyceps sinensis, contribute to the inhibition of the development of renal cysts and the progression of ADPKD, which function by similar or different mechanisms. These studies suggest that herbal medicines could be a promising type of drugs and can provide new inspiration for clinical therapeutic strategy for ADPKD. This review summarizes the pharmacological effects of the herbal medicines on ADPKD progression and their underlying mechanisms in both in vivo and in vitro ADPKD models.

scholarly journals Discovery and preclinical evaluation of anti-miR-17 oligonucleotide RGLS4326 for the treatment of polycystic kidney disease

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Edmund C. Lee ◽  
Tania Valencia ◽  
Charles Allerson ◽  
Annelie Schairer ◽  
Andrea Flaten ◽  
...  
Keyword(s):  
Kidney Disease ◽  
Polycystic Kidney Disease ◽  
Collecting Duct ◽  
Treatment Options ◽  
Subcutaneous Administration ◽  
Polycystic Kidney ◽  
Short Oligonucleotide ◽  
Mrna Targets ◽  
Stage Renal Disease

Abstract Autosomal dominant polycystic kidney disease (ADPKD), caused by mutations in either PKD1 or PKD2 genes, is one of the most common human monogenetic disorders and the leading genetic cause of end-stage renal disease. Unfortunately, treatment options for ADPKD are limited. Here we report the discovery and characterization of RGLS4326, a first-in-class, short oligonucleotide inhibitor of microRNA-17 (miR-17), as a potential treatment for ADPKD. RGLS4326 is discovered by screening a chemically diverse and rationally designed library of anti-miR-17 oligonucleotides for optimal pharmaceutical properties. RGLS4326 preferentially distributes to kidney and collecting duct-derived cysts, displaces miR-17 from translationally active polysomes, and de-represses multiple miR-17 mRNA targets including Pkd1 and Pkd2. Importantly, RGLS4326 demonstrates a favorable preclinical safety profile and attenuates cyst growth in human in vitro ADPKD models and multiple PKD mouse models after subcutaneous administration. The preclinical characteristics of RGLS4326 support its clinical development as a disease-modifying treatment for ADPKD.

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