Mouse Model Of Autosomal Dominant Polycystic Kidney Disease Research Articles

Circadian clock disruption and growth of kidney cysts in autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in the PKD1 and PKD2 genes, and often progresses to kidney failure. ADPKD progression is not uniform among patients, suggesting that factors secondary to the PKD1/2 gene mutation could regulate the rate of disease progression. Here we tested the effect of circadian clock disruption on ADPKD progression. Circadian rhythms are regulated by cell-autonomous circadian clocks composed of clock proteins. BMAL1 is a core constituent of the circadian clock. To disrupt the circadian clock, we deleted Bmal1 gene in the renal collecting ducts of the Pkd1 RC/RC (RC/RC) mouse model of ADPKD (RC/RC; Bmal1 f/f ; Pkhd1 cre , called DKO mice), and in Pkd1 knockout mouse inner medullary collecting duct cells ( Pkd1Bmal1 KO mIMCD3 cells). Only male mice were used. Human nephrectomy ADPKD kidneys and Pkd1 KO mIMCD3 cells showed reduced Bmal1 gene expression compared to normal controls. When compared to RC/RC kidneys, DKO kidneys showed significantly altered clock gene expression, increased cyst growth, cell proliferation, apoptosis and fibrosis. DKO kidneys also showed increased lipogenesis and cholesterol synthesis-related gene expression, and increased tissue triglyceride levels compared to RC/RC kidneys. Similarly, in vitro, Pkd1Bmal1 KO cells showed altered clock genes, increased lipogenesis and cholesterol synthesis-related genes, and reduced fatty-acid oxidation-related gene expression compared to Pkd1KO cells. The Pkd1Bmal1 KO cells showed increased cell proliferation compared to Pkd1KO cells, which was rescued by pharmacological inhibition of lipogenesis. Renal collecting duct specific Bmal1 gene deletion disrupts the circadian clock and triggers accelerated ADPKD progression by altering lipid metabolism-related gene expression. Lack of BMAL1, a circadian clock protein in renal collecting ducts disrupted the clock and increased cyst growth and fibrosis in an ADPKD mouse model.BMAL1 gene deletion increased cell proliferation by increasing lipogenesis in kidney cells.Thus, circadian clock disruption could be a risk factor for accelerated disease progression in patients with ADPKD.

HL156A, an AMP-Activated Protein Kinase Activator, Inhibits Cyst Growth in Autosomal Dominant Polycystic Kidney Disease.

Autosomal dominant polycystic kidney disease (ADPKD) is the most prevalent genetic kidney disorder. While metformin has demonstrated the ability to inhibit cyst growth in animal models of ADPKD via activation of adenosine monophosphate-activated protein kinase (AMPK), its effectiveness in humans is limited due to its low potency. This study explored the impact of HL156A, a new and more potent AMPK activator, in a mouse model of ADPKD. To investigate whether HL156A inhibits the proliferation of renal cyst cells in ADPKD in vitro, exogenous human telomerase reverse transcriptase (hTERT)-immortalized renal cyst cells from ADPKD patients were treated with HL156A, and an MTT (dimethylthiazol-diphenyltetrazolium bromide) assay was performed. To assess the cyst-inhibitory effect of HL156A in vivo, we generated Pkd1 conditional knockout (KO) mice with aquaporin 2 (AQP2)-Cre, which selectively expresses Cre recombinase in the collecting duct. The effectiveness of HL156A in inhibiting cyst growth and improving renal function was confirmed by measuring the number of cysts and blood urea nitrogen (BUN) levels in the collecting duct-specific Pkd1 KO mice. When cyst cells were treated with up to 20 µM of metformin or HL156A, HL156A reduced cell viability by 25% starting at a concentration of 5 µM, whereas metformin showed no effect. When AQP2-Cre male mice were crossed with Pkd1flox/flox female mice, and when AQP2-Cre female mice were crossed with Pkd1flox/flox male mice, the number of litters produced by both groups was comparable. In collecting duct-specific Pkd1 KO mice, HL156A was found to inhibit cyst growth, reducing both the number and size of cysts. Furthermore, it was confirmed that kidney function improved as HL156A treatment led to a reduction in elevated BUN levels. Lastly, it was observed that the increase in AMPK phosphorylation induced by HL156A decreased ERK phosphorylation and α-SMA expression. HL156A has potential as a drug that can restore kidney function in ADPKD patients by inhibiting cyst growth.

P2Y2R and Cyst Growth in Polycystic Kidney Disease.

Key Points Polycystic kidney disease (PKD) is characterized by continuous cyst growth, which results in a decline in kidney function.Deletion of P2Y2R and pharmacological antagonism of purinergic signaling significantly reduced cyst growth in an orthologous PKD mouse model.P2Y2R was expressed in cysts of human PKD nephrectomies, which makes P2Y2R a reasonable target for treatment of PKD. Background Autosomal dominant polycystic kidney disease (ADPKD) is characterized by multiple bilateral kidney cysts that gradually enlarge, resulting in a decline in kidney function. Cyst growth is significantly driven by ATP-dependent chloride secretion mediated by the ion channel TMEM16A. This pathway is further augmented in advanced stages of the disease by hypoxia and activation of hypoxia-inducible factor (HIF)-1α. The mechanisms by which ATP leads to activation of TMEM16A and how HIF-1α contributes to cyst growth in vivo have remained elusive. Methods Mice with an inducible tubule-specific deletion of Pkd1 were compared with mice with an additional codeletion of the purinergic receptor P2y2r. Furthermore, animals were challenged by pharmacological activation of HIF-1α and Pkd1-deficient mice were treated with suramin, an antagonist of purinergic signaling. In addition, expression of P2Y2R, TMEM16A, and HIF-1α was analyzed in nephrectomy samples from 27 patients with ADPKD. Results Genetic deletion of P2y2r significantly inhibited cyst growth in vivo. In addition, aggravation of the polycystic phenotype mediated by pharmacological activation of HIF-1α was reduced by deletion of P2y2r. Application of suramin to pharmacologically inhibit purinergic signaling also suppressed cyst enlargement in vivo. Analysis of kidney samples from 27 patients with ADPKD revealed significant expression of P2Y2R at the luminal site of the cyst-lining epithelium. Conclusions P2Y2R was significantly expressed in human and mouse polycystic kidneys. Deletion and antagonism of P2Y2R reduced cyst enlargement in an ADPKD mouse model.

Glis2 is an early effector of polycystin signaling and a target for therapy in polycystic kidney disease

Mouse models of autosomal dominant polycystic kidney disease (ADPKD) show that intact primary cilia are required for cyst growth following the inactivation of polycystin-1. The signaling pathways underlying this process, termed cilia-dependent cyst activation (CDCA), remain unknown. Using translating ribosome affinity purification RNASeq on mouse kidneys with polycystin-1 and cilia inactivation before cyst formation, we identify the differential ‘CDCA pattern’ translatome specifically dysregulated in kidney tubule cells destined to form cysts. From this, Glis2 emerges as a candidate functional effector of polycystin signaling and CDCA. In vitro changes in Glis2 expression mirror the polycystin- and cilia-dependent changes observed in kidney tissue, validating Glis2 as a cell culture-based indicator of polycystin function related to cyst formation. Inactivation of Glis2 suppresses polycystic kidney disease in mouse models of ADPKD, and pharmacological targeting of Glis2 with antisense oligonucleotides slows disease progression. Glis2 transcript and protein is a functional target of CDCA and a potential therapeutic target for treating ADPKD.

High-Resolution Spatial Lipidomic Profiling of the Kidney During Periods of Cyst Expansion in a Mouse Model of ADPKD

Introduction: The progression of renal pathology in autosomal dominant polycystic kidney disease (ADPKD) involves a complex series of changes including renal cyst formation, interstitial cyst expansion, tubular atrophy, and fibrosis. Changes to in tissue metabolism and cyst fluid metabolite composition have been shown in ADPKD models, however it has been diffcult to discretely localize these compounds to specific regions of the kidney. Further, very little is known about lipid profiles within the kidney during ADPKD. We hypothesized that spatial lipidomic analysis during periods of cyst expansion would reveal new understanding of changes in lipid metabolism during this critical window of pathology. In this study we focused on changes in the mcwPkd1nl/nl mouse, a hypomorphic ADPKD model that rapidly develops a cystic phenotype. Methods: Kidneys were collected from female cystic mcwPkd1nl/nl mice at postnatal days 7, 14 and 28, representing periods preceding, during and at maximal cyst expansion, respectively. Control kidneys were collected from female wild-type littermates at postnatal day 7 and 28. Tissues were rapidly cryoembedded for 10 mm sectioning onto Intellislides (BrukerDaltonik). Slides were scanned on a Reflecta MF-5000 scanner and sprayed with 7 mg/mL CHCA in 70% Acetonitrile, 30% water and 0.1% TFA using a TM Sprayer Model M3 (HTX Technologies, LLC). Mass spec imaging data was acquired with a Bruker timsTOF Flex system (BrukerDaltonik) with a Smartbeam 3D laser at 10kHz and 10 μm resolution. Imaging data analysis was performed in SCiLS Lab software (version 2023b, Bruker). Calculations were performed on data normalized to total ion count (TIC). Spatial segmentation analysis of data was performed using an unsupervised bisecting k-means algorithm and resulting clusters were investigated by ROC. Lipid annotation was performed the Lipid Species annotation tool with MSDial VS68 spectral library in Metaboscape (version 2022b, Bruker). Results: There were vast global differences between lipid composition of wild-type and cystic kidneys at postnatal day 28. When comparing cystic to wild-type kidneys phosphoinositol and ceremide lipid species were decreased and other metabolites, such as thiophene-3-carboxylate and benzothiazine-2-carbonitrile, were increased at both PN7 and PN28. Further numerous lipid compounds were identified only within cystic fluid that have not been previously annotated. Conclusions: The results of this study, for the first time, reveals alterations in lipid molecule profiles throughout the cystic kidney in tandem with morphological changes. This is a major step forward in understanding how lipid metabolism changes in response to cyst expansion during the progression of ADPKD, as well as revealing molecular changes that precede cellular pathology. This work has been supported by Children’s Wisconsin Foundation and Children’s Research Institute. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Benzothiazole derivatives as histone deacetylase inhibitors for the treatment of autosomal dominant polycystic kidney disease

Recent evidence suggests that histone deacetylases (HDACs) are important regulators of autosomal dominant polycystic kidney disease (ADPKD). In the present study, a series of benzothiazole-bearing compounds were designed and synthesized as potential HDAC inhibitors. Given the multiple participation of HDACs in ADPKD cyst progression, we embarked on a targeted screen using HeLa nuclear extracts to identify potent pan-HDAC inhibitors. Compound 26 emerged as the most efficacious candidate. Subsequent pharmacological characterization showed that compound 26 effectively inhibits several HDACs, notably HDAC1, HDAC2, and HDAC6 (IC50 < 150 nM), displaying a particularly high sensitivity towards HDAC6 (IC50 = 11 nM). The selected compound significantly prevented cyst formation and expansion in an in vitro cyst model and was efficacious in reducing cyst growth in both an embryonic kidney cyst model and an in vivo ADPKD mouse model. Our results provided compelling evidence that compound 26 represents a new HDAC inhibitor for the treatment of ADPKD.

Inhibiting centrosome clustering reduces cystogenesis and improves kidney function in autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is a monogenic disorder accounting for approximately 5% of patients with renal failure, yet therapeutics for the treatment of ADPKD remain limited. ADPKD tissues display abnormalities in the biogenesis of the centrosome, a defect that can cause genome instability, aberrant ciliary signaling, and secretion of pro-inflammatory factors. Cystic cells form excess centrosomes via a process termed centrosome amplification (CA), which causes abnormal multipolar spindle configurations, mitotic catastrophe, and reduced cell viability. However, cells with CA can suppress multipolarity via "centrosome clustering," a key mechanism by which cells circumvent apoptosis. Here, we demonstrate that inhibiting centrosome clustering can counteract the proliferation of renal cystic cells with high incidences of CA. Using ADPKD human cells and mouse models, we show that preventing centrosome clustering with 2 inhibitors, CCB02 and PJ34, blocks cyst initiation and growth in vitro and in vivo. Inhibiting centrosome clustering activates a p53-mediated surveillance mechanism leading to apoptosis, reduced cyst expansion, decreased interstitial fibrosis, and improved kidney function. Transcriptional analysis of kidneys from treated mice identified pro-inflammatory signaling pathways implicated in CA-mediated cystogenesis and fibrosis. Our results demonstrate that centrosome clustering is a cyst-selective target for the improvement of renal morphology and function in ADPKD.

A synthetic agent ameliorates polycystic kidney disease by promoting apoptosis of cystic cells through increased oxidative stress

Autosomal dominant polycystic kidney disease (ADPKD) is the most common monogenic cause of chronic kidney disease and the fourth leading cause of end-stage kidney disease, accounting for over 50% of prevalent cases requiring renal replacement therapy. There is a pressing need for improved therapy for ADPKD. Recent insights into the pathophysiology of ADPKD revealed that cyst cells undergo metabolic changes that up-regulate aerobic glycolysis in lieu of mitochondrial respiration for energy production, a process that ostensibly fuels their increased proliferation. The present work leverages this metabolic disruption as a way to selectively target cyst cells for apoptosis. This small-molecule therapeutic strategy utilizes 11beta-dichloro, a repurposed DNA-damaging anti-tumor agent that induces apoptosis by exacerbating mitochondrial oxidative stress. Here, we demonstrate that 11beta-dichloro is effective in delaying cyst growth and its associated inflammatory and fibrotic events, thus preserving kidney function in perinatal and adult mouse models of ADPKD. In both models, the cyst cells with homozygous inactivation of Pkd1 show enhanced oxidative stress following treatment with 11beta-dichloro and undergo apoptosis. Co-administration of the antioxidant vitamin E negated the therapeutic benefit of 11beta-dichloro in vivo, supporting the conclusion that oxidative stress is a key component of the mechanism of action. As a preclinical development primer, we also synthesized and tested an 11beta-dichloro derivative that cannot directly alkylate DNA, while retaining pro-oxidant features. This derivative nonetheless maintains excellent anti-cystic properties in vivo and emerges as the lead candidate for development.

Human iPSC-derived renal collecting duct organoid model cystogenesis in ADPKD

In autosomal dominant polycystic kidney disease (ADPKD), renal cyst lesions predominantly arise from collecting ducts (CDs). However, relevant CD cyst models using human cells are lacking. Although previous reports have generated invitro renal tubule cyst models from human induced pluripotent stem cells (hiPSCs), therapeutic drug candidates for ADPKD have not been identified. Here, by establishing expansion cultures of hiPSC-derived ureteric bud tip cells, an embryonic precursor that gives rise to CDs, we succeed in advancing the developmental stage of CD organoids and show that all CD organoids derived from PKD1-/- hiPSCs spontaneously develop multiple cysts, clarifying the initiation mechanisms of cystogenesis. Moreover, we identify retinoic acid receptor (RAR) agonists as candidate drugs that suppress invitro cystogenesis and confirm the therapeutic effects on an ADPKD mouse model invivo. Therefore, our invitro CD cyst model contributes to understanding disease mechanisms and drug discovery for ADPKD.

In vitro delivery of mTOR inhibitors by kidney-targeted micelles for autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is the most common genetic kidney disease and is characterized by the formation of renal cysts and the eventual development of end-stage kidney disease. One approach to treating ADPKD is through inhibition of the mammalian target of rapamycin (mTOR) pathway, which has been implicated in cell overproliferation, contributing to renal cyst expansion. However, mTOR inhibitors, including rapamycin, everolimus, and RapaLink-1, have off-target side effects including immunosuppression. Thus, we hypothesized that the encapsulation of mTOR inhibitors in drug delivery carriers that target the kidneys would provide a strategy that would enable therapeutic efficacy while minimizing off-target accumulation and associated toxicity. Toward eventual in vivo application, we synthesized cortical collecting duct (CCD) targeted peptide amphiphile micelle (PAM) nanoparticles and show high drug encapsulation efficiency (>92.6%). In vitro analysis indicated that drug encapsulation into PAMs enhanced the anti-proliferative effect of all three drugs in human CCD cells. Analysis of in vitro biomarkers of the mTOR pathway via western blotting confirmed that PAM encapsulation of mTOR inhibitors did not reduce their efficacy. These results indicate that PAM encapsulation is a promising way to deliver mTOR inhibitors to CCD cells and potentially treat ADPKD. Future studies will evaluate the therapeutic effect of PAM-drug formulations and ability to prevent off-target side effects associated with mTOR inhibitors in mouse models of ADPKD.

A novel direct adenosine monophosphate kinase activator ameliorates disease progression in preclinical models of Autosomal Dominant Polycystic Kidney Disease

Autosomal dominant polycystic kidney disease (ADPKD) mainly results from mutations in the PKD1 gene which encodes polycystin 1. It is the most common inherited kidney disease and is characterized by a progressive bilateral increase in cyst number and size, often leading to kidney failure. The cellular energy sensor and regulator, adenosine monophosphate stimulated protein kinase (AMPK), has been implicated as a promising new therapeutic target. To address this hypothesis, we determined the effects of a potent and selective clinical stage direct allosteric AMPK activator, PXL770, in canine and patient-derived 3D cyst models and an orthologous mouse model of ADPKD. PXL770 induced AMPK activation and dose-dependently reduced cyst growth in principal-like Madin-Darby Canine Kidney cells stimulated with forskolin and kidney epithelial cells derived from patients with ADPKD stimulated with desmopressin. In an inducible, kidney epithelium-specific Pkd1 knockout mouse model, PXL770 produced kidney AMPK pathway engagement, prevented the onset of kidney failure (reducing blood urea by 47%), decreased cystic index by 26% and lowered the kidney weight to body weight ratio by 35% compared to untreated control Pkd1 knockout mice. These effects were accompanied by a reduction of markers of cell proliferation (-48%), macrophage infiltration (-53%) and tissue fibrosis (-37%). Thus, our results show the potential of direct allosteric AMPK activation in the treatment of ADPKD and support the further development of PXL770 for this indication.

Discovery of Novel N-(5-(Pyridin-3-yl)-1H-indazol-3-yl)benzamide Derivatives as Potent Cyclin-Dependent Kinase 7 Inhibitors for the Treatment of Autosomal Dominant Polycystic Kidney Disease.

Recent evidence suggests that CDK7 is a novel potential drug target for autosomal dominant polycystic kidney disease (ADPKD) treatment. Herein, on the basis of structural analysis, a hit compound 3 with a novel scaffold was designed and subsequent medicinal chemistry efforts by a rational design strategy were conducted to improve CDK7 inhibitors' potency and selectivity. The representative compound B2 potently inhibited CDK7 with an IC50 value of 4 nM and showed high selectivity over CDKs. Compound B2 showed high potency to inhibit cyst growth and exhibited lower cytotoxicity than THZ1 in an in vitro Madin-Darby canine kidney cyst model. In addition, compound B2 was also highly efficacious in suppressing renal cyst development in an ex vivo embryonic kidney cyst model and in vivo ADPKD mouse model. These results indicate that compound B2 represents a promising lead compound that deserves further investigation to discover novel therapeutic agents for ADPKD.

Myofibroblast depletion reduces kidney cyst growth and fibrosis in autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) involves the development and persistent growth of fluid filled kidney cysts. In a recent study, we showed that ADPKD kidney cyst epithelial cells can stimulate the proliferation and differentiation of peri-cystic myofibroblasts. Although dense myofibroblast populations are often found surrounding kidney cysts, their role in cyst enlargement or fibrosis in ADPKD is unclear. To clarify this, we examined the effect of myofibroblast depletion in the Pkd1RC/RC (RC/RC) mouse model of ADPKD. RC/RC;αSMAtk mice that use the ganciclovir-thymidine kinase system to selectively deplete α-smooth muscle actin expressing myofibroblasts were generated. Ganciclovir treatment for four weeks depleted myofibroblasts, reduced kidney fibrosis and preserved kidney function in these mice. Importantly, myofibroblast depletion significantly reduced cyst growth and cyst epithelial cell proliferation in RC/RC;αSMAtk mouse kidneys. Similar ganciclovir treatment did not alter cyst growth or fibrosis in wild-type or RC/RC littermates. Invitro, co-culture with myofibroblasts from the kidneys of patients with ADPKD increased 3D microcyst growth of human ADPKD cyst epithelial cells. Treatment with conditioned culture media from ADPKD kidney myofibroblasts increased microcyst growth and cell proliferation of ADPKD cyst epithelial cells. Further examination of ADPKD myofibroblast conditioned media showed high levels of protease inhibitors including PAI1, TIMP1 and 2, NGAL and TFPI-2, and treatment with recombinant PAI1 and TIMP1 increased ADPKD cyst epithelial cell proliferation invitro. Thus, our findings show that myofibroblasts directly promote cyst epithelial cell proliferation, cyst growth and fibrosis in ADPKD kidneys, and their targeting could be a novel therapeutic strategy to treat PKD.

1-Indanone retards cyst development in ADPKD mouse model by stabilizing tubulin and down-regulating anterograde transport of cilia.

Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disease. Cyst development in ADPKD involves abnormal epithelial cell proliferation, which is affected by the primary cilia-mediated signal transduction in the epithelial cells. Thus, primary cilium has been considered as a therapeutic target for ADPKD. Since ADPKD exhibits many pathological features similar to solid tumors, we investigated whether targeting primary cilia using anti-tumor agents could alleviate the development of ADPKD. Twenty-four natural compounds with anti-tumor activity were screened in MDCK cyst model, and 1-Indanone displayed notable inhibition on renal cyst growth without cytotoxicity. This compound also inhibited cyst development in embryonic kidney cyst model. In neonatal kidney-specific Pkd1 knockout mice, 1-Indanone remarkably slowed down kidney enlargement and cyst expansion. Furthermore, we demonstrated that 1-Indanone inhibited the abnormal elongation of cystic epithelial cilia by promoting tubulin polymerization and significantly down-regulating expression of anterograde transport motor protein KIF3A and IFT88. Moreover, we found that 1-Indanone significantly down-regulated ciliary coordinated Wnt/β-catenin, Hedgehog signaling pathways. These results demonstrate that 1-Indanone inhibits cystic cell proliferation by reducing abnormally prolonged cilia length in cystic epithelial cells, suggesting that 1-Indanone may hold therapeutic potential to retard cyst development in ADPKD.

PF-06409577 inhibits renal cyst progression by concurrently inhibiting the mTOR pathway and CFTR channel activity.

Renal cyst development and expansion in autosomal dominant polycystic kidney disease (ADPKD) involves over‐proliferation of cyst‐lining epithelial cells and excessive cystic fluid secretion. While metformin effectively inhibits renal cyst growth in mouse models of ADPKD it exhibits low potency, and thus an adenosine monophosphate‐activated protein kinase (AMPK) activator with higher potency is required. Herein, we adopted a drug repurposing strategy to explore the potential of PF‐06409577, an AMPK activator for diabetic nephropathy, in cellular, ex vivo and in vivo models of ADPKD. Our results demonstrated that PF‐06409577 effectively down‐regulated mammalian target of rapamycin pathway‐mediated proliferation of cyst‐lining epithelial cells and reduced cystic fibrosis transmembrane conductance regulator‐regulated cystic fluid secretion. Overall, our data suggest that PF‐06409577 holds therapeutic potential for ADPKD treatment.

CAMP-Induced Nuclear Condensation of CRTC2 Promotes Transcription Elongation and Cystogenesis in Autosomal Dominant Polycystic Kidney Disease.

Formation of biomolecular condensates by phase separation has recently emerged as a new principle for regulating gene expression in response to extracellular signaling. However, the molecular mechanisms underlying the coupling of signal transduction and gene activation through condensate formation, and how dysregulation of these mechanisms contributes to disease progression, remain elusive. Here, the authors report that CREB‐regulated transcription coactivator 2 (CRTC2) translocates to the nucleus and forms phase‐separated condensates upon activation of cAMP signaling. They show that intranuclear CRTC2 interacts with positive transcription elongation factor b (P‐TEFb) and activates P‐TEFb by disrupting the inhibitory 7SK snRNP complex. Aberrantly elevated cAMP signaling plays central roles in the development of autosomal dominant polycystic kidney disease (ADPKD). They find that CRTC2 localizes to the nucleus and forms condensates in cystic epithelial cells of both mouse and human ADPKD kidneys. Genetic depletion of CRTC2 suppresses cyst growth in an orthologous ADPKD mouse model. Using integrative transcriptomic and cistromic analyses, they identify CRTC2‐regulated cystogenesis‐associated genes, whose activation depends on CRTC2 condensate‐facilitated P‐TEFb recruitment and the release of paused RNA polymerase II. Together, their findings elucidate a mechanism by which CRTC2 nuclear condensation conveys cAMP signaling to transcription elongation activation and thereby promotes cystogenesis in ADPKD.

Weight loss and cystic disease progression in autosomal dominant polycystic kidney disease

SummaryProgression of autosomal dominant polycystic kidney disease (ADPKD) is modified by metabolic defects and obesity. Indeed, reduced food intake slows cyst growth in preclinical rodent studies. Here, we demonstrate the feasibility of daily caloric restriction (DCR) and intermittent fasting (IMF) in a cohort of overweight or obese patients with ADPKD. Clinically significant weight loss occurred with both DCR and IMF; however, weight loss was greater and adherence and tolerability were better with DCR. Further, slowed kidney growth correlated with body weight and visceral adiposity loss independent of dietary regimen. Similarly, we compared the therapeutic efficacy of DCR, IMF, and time restricted feeding (TRF) using an orthologous ADPKD mouse model. Only ADPKD animals on DCR lost significant weight and showed slowed cyst growth compared to ad libitum, IMF, or TRF feeding. Collectively, this supports therapeutic feasibility of caloric restriction in ADPKD, with potential efficacy benefits driven by weight loss.

The Mitochondrial Ca2+ import complex is altered in ADPKD

Mutations in either of the polycystic kidney disease genes, PKD1 or PKD2, engender the growth of cysts, altering renal function. Cystic growth is supported by major changes in cellular metabolism, some of which involve the mitochondrion, a major storage site for Ca2+ and a key organelle in cellular Ca2+ signaling. The goal here was to understand the role of components of the mitochondrial Ca2+ uptake complex in PC1-mutant cells in autosomal dominant polycystic kidney disease (ADPKD). We found that the mitochondrial Ca2+ uniporter (MCU) and voltage-dependent anion channels 1& 3 (VDAC) were down-regulated in different mouse and cell models of ADPKD along with the Ca2+-dependent enzyme, pyruvate dehydrogenase phosphatase (PDHX). The release of Ca2+ from the endoplasmic reticulum, and Ca2+ uptake by the mitochondria were upregulated in PC1(polycystin)-null cells. We also observed an enhanced staining with MitoTracker Red CMXRos in PC1-null cultured cells than in PC1-containing cells and a substantially higher increase in response to ER Ca2+ release. Increased colocalization of the Ca2+ sensitive dye, rhodamine2, with MitoTracker Green suggested an increase Ca2+ entry into the mitochondria in PC1 null cells subsequent to Ca2+ release from the ER or from Ca2+ entry from the extracellular solution. These data clearly demonstrate abnormal release of Ca2+ by the ER and corresponding alterations in Ca2+ uptake by the mitochondria in PC1−null cells. Importantly, inhibiting mitochondrial Ca2+ uptake with the specific inhibitor Ru360 inhibited cyst growth and altered both apoptosis and cell proliferation. We further show that the decrease in mitochondrial proteins and abnormally high Ca2+ signaling can be reversed by application of the cystic fibrosis (CFTR) corrector, VX-809. We conclude that enhanced Ca2+ signaling and alterations in proteins association with the mitochondrial Ca2+ uptake complex are associated with malfunction of PC1. Finally, our results identify novel therapeutic targets for treating ADPKD.

β3 adrenergic receptor as potential therapeutic target in ADPKD.

Autosomal dominant polycystic kidney disease (ADPKD) disrupts renal parenchyma through progressive expansion of fluid‐filled cysts. The only approved pharmacotherapy for ADKPD involves the blockade of the vasopressin type 2 receptor (V2R). V2R is a GPCR expressed by a subset of renal tubular cells and whose activation stimulates cyclic AMP (cAMP) accumulation, which is a major driver of cyst growth. The β3‐adrenergic receptor (β3‐AR) is a GPCR expressed in most segments of the murine nephron, where it modulates cAMP production. Since sympathetic nerve activity, which leads to activation of the β3‐AR, is elevated in patients affected by ADPKD, we hypothesize that β3‐AR might constitute a novel therapeutic target. We find that administration of the selective β3‐AR antagonist SR59230A to an ADPKD mouse model (Pkd1fl/fl;Pax8rtTA;TetO‐Cre) decreases cAMP levels, producing a significant reduction in kidney/body weight ratio and a partial improvement in kidney function. Furthermore, cystic mice show significantly higher β3‐AR levels than healthy controls, suggesting a correlation between receptor expression and disease development. Finally, β3‐AR is expressed in human renal tissue and localizes to cyst‐lining epithelial cells in patients. Thus, β3‐AR is a potentially interesting target for the development of new treatments for ADPKD.

The tyrosine-kinase inhibitor Nintedanib ameliorates autosomal-dominant polycystic kidney disease

Autosomal-dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disease and is characterized by progressive growth of fluid-filled cysts. Growth factors binding to receptor tyrosine kinases (RTKs) stimulate cell proliferation and cyst growth in PKD. Nintedanib, a triple RTK inhibitor, targets the vascular endothelial growth-factor receptor (VEGFR), platelet-derived growth-factor receptor (PDGFR), and fibroblast growth-factor receptor (FGFR), and is an approved drug for the treatment of non-small-cell lung carcinoma and idiopathic lung fibrosis. To determine if RTK inhibition using nintedanib can slow ADPKD progression, we tested its effect on human ADPKD renal cyst epithelial cells and myofibroblasts in vitro, and on Pkd1f/fPkhd1Cre and Pkd1RC/RC, orthologous mouse models of ADPKD. Nintedanib significantly inhibited cell proliferation and in vitro cyst growth of human ADPKD renal cyst epithelial cells, and cell viability and migration of human ADPKD renal myofibroblasts. Consistently, nintedanib treatment significantly reduced kidney-to-body-weight ratio, renal cystic index, cystic epithelial cell proliferation, and blood-urea nitrogen levels in both the Pkd1f/fPkhd1Cre and Pkd1RC/RC mice. There was a corresponding reduction in ERK, AKT, STAT3, and mTOR activity and expression of proproliferative factors, including Yes-associated protein (YAP), c-Myc, and Cyclin D1. Nintedanib treatment significantly reduced fibrosis in Pkd1RC/RC mice, but did not affect renal fibrosis in Pkd1f/fPkhd1Cre mice. Overall, these results suggest that nintedanib may be repurposed to effectively slow cyst growth in ADPKD.