The Structure of the Polycystic Kidney Disease Channel PKD2 in Lipid Nanodiscs.

The Polycystic Kidney Disease 2 (Pkd2) gene is mutated in autosomal dominant polycystic kidney disease (ADPKD), one of the most common human monogenic disorders. Here, we present the cryo-EM structure of PKD2 in lipid bilayers at 3.0 Å resolution, which establishes PKD2 as a homotetrameric ion channel and provides insight into potential mechanisms for its activation. The PKD2 voltage-sensor domain retains two of four gating charges commonly found in those of voltage-gated ion channels. The PKD2 ion permeation pathway is constricted at the selectivity filter and near the cytoplasmic end of S6, suggesting that two gates regulate ion conduction. The extracellular domain of PKD2, a hotspot for ADPKD pathogenic mutations, contributes to channel assembly and strategically interacts with the transmembrane core, likely serving as a physical substrate for extracellular stimuli to allosterically gate the channel. Finally, our structure establishes the molecular basis for the majority of pathogenic mutations in Pkd2-related ADPKD.

Polycystin Channel Complexes.

Polycystin subunits can form hetero- and homotetrameric ion channels in the membranes of various compartments of the cell. Homotetrameric polycystin channels are voltage- and calcium-modulated, whereas heterotetrameric versions are proposed to be ligand- or autoproteolytically regulated. Their importance is underscored by variants associated with autosomal dominant polycystic kidney disease and by vital roles in fertilization and embryonic development. The diversity in polycystin assembly and subcellular distribution allows for a multitude of sensory functions by this class of channels. In this review, we highlight their recent structural and functional characterization, which has provided a molecular blueprint to investigate the conformational changes required for channel opening in response to unique stimuli. We consider each polycystin channel type individually, discussing how they contribute to sensory cell biology, as well as their impact on the physiology of various tissues.

Polycystic kidney disease: a Hippo connection.

Mutations in PKD1 and PKD2 are the leading cause of autosomal dominant polycystic kidney disease (ADPKD). In this issue of Genes & Development, a report by Cai and colleagues (pp. 781-793) reveals new insight into the molecular basis by which PKD1 deficiency leads to cystic kidney pathogenesis. By using extensive mouse genetic analyses coupled with in vitro cystic assays, the investigators delineate a RhoA-YAP-c-Myc signaling axis as a key downstream from PKD1 deficiency in ADPKD pathogenesis. Their findings provide evidence that the Hippo pathway could be a potential target for treating ADPKD.

A RhoA-YAP-c-Myc signaling axis promotes the development of polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is an inherited disorder caused by mutations in PKD1 or PKD2 and affects one in 500-1000 humans. Limited treatment is currently available for ADPKD. Here we identify the Hippo signaling effector YAP and its transcriptional target, c-Myc, as promoters of cystic kidney pathogenesis. While transgenic overexpression of YAP promotes proliferation and tubule dilation in mouse kidneys, loss of YAP/TAZ or c-Myc suppresses cystogenesis in a mouse ADPKD model resulting from Pkd1 deficiency. Through a comprehensive kinase inhibitor screen based on a novel three-dimensional (3D) culture of Pkd1 mutant mouse kidney cells, we identified a signaling pathway involving the RhoGEF (guanine nucleotide exchange factor) LARG, the small GTPase RhoA, and the RhoA effector Rho-associated kinase (ROCK) as a critical signaling module between PKD1 and YAP. Further corroborating its physiological importance, inhibition of RhoA signaling suppresses cystogenesis in 3D culture of Pkd1 mutant kidney cells as well as Pkd1 mutant mouse kidneys in vivo. Taken together, our findings implicate the RhoA-YAP-c-Myc signaling axis as a critical mediator and potential drug target in ADPKD.

Disease-associated missense mutations in the pore loop of polycystin-2 alter its ion channel function in a heterologous expression system.

Polycystin-2 (PC2) is mutated in ∼15% of patients with autosomal dominant polycystic kidney disease (ADPKD). PC2 belongs to the family of transient receptor potential (TRP) channels and can function as a homotetramer. We investigated whether three disease-associated mutations (F629S, C632R, or R638C) localized in the channel's pore loop alter ion channel properties of human PC2 expressed in Xenopus laevis oocytes. Expression of wild-type (WT) PC2 typically resulted in small but measurable Na+ inward currents in the absence of extracellular divalent cations. These currents were no longer observed when individual pore mutations were introduced in WT PC2. Similarly, Na+ inward currents mediated by the F604P gain-of-function (GOF) PC2 construct (PC2 F604P) were abolished by each of the three pore mutations. In contrast, when the mutations were introduced in another GOF construct, PC2 L677A N681A, only C632R had a complete loss-of-function effect, whereas significant residual Na+ inward currents were observed with F629S (∼15%) and R638C (∼30%). Importantly, the R638C mutation also abolished the Ca2+ permeability of PC2 L677A N681A and altered its monovalent cation selectivity. To elucidate the molecular mechanisms by which the R638C mutation affects channel function, molecular dynamics (MD) simulations were used in combination with functional experiments and site-directed mutagenesis. Our findings suggest that R638C stabilizes ionic interactions between Na+ ions and the selectivity filter residue D643. This probably explains the reduced monovalent cation conductance of the mutant channel. In summary, our data support the concept that altered ion channel properties of PC2 contribute to the pathogenesis of ADPKD.

Monoallelic IFT140 pathogenic variants are an important cause of the autosomal dominant polycystic kidney-spectrum phenotype.

Autosomal dominant polycystic kidney disease (ADPKD), characterized by progressive cyst formation/expansion, results in enlarged kidneys and often end stage kidney disease. ADPKD is genetically heterogeneous; PKD1 and PKD2 are the common loci (∼78% and ∼15% of families) and GANAB, DNAJB11, and ALG9 are minor genes. PKD is a ciliary-associated disease, a ciliopathy, and many syndromic ciliopathies have a PKD phenotype. In a multi-cohort/-site collaboration, we screened ADPKD-diagnosed families that were naive to genetic testing (n = 834) or for whom no PKD1 and PKD2 pathogenic variants had been identified (n = 381) with a PKD targeted next-generation sequencing panel (tNGS; n = 1,186) or whole-exome sequencing (WES; n = 29). We identified monoallelic IFT140 loss-of-function (LoF) variants in 12 multiplex families and 26 singletons (1.9% of naive families). IFT140 is a core component of the intraflagellar transport-complex A, responsible for retrograde ciliary trafficking and ciliary entry of membrane proteins; bi-allelic IFT140 variants cause the syndromic ciliopathy, short-rib thoracic dysplasia (SRTD9). The distinctive monoallelic phenotype is mild PKD with large cysts, limited kidney insufficiency, and few liver cysts. Analyses of the cystic kidney disease probands of Genomics England 100K showed that 2.1% had IFT140 LoF variants. Analysis of the UK Biobank cystic kidney disease group showed probands with IFT140 LoF variants as the third most common group, after PKD1 and PKD2. The proximity of IFT140 to PKD1 (∼0.5 Mb) in 16p13.3 can cause diagnostic confusion, and PKD1 variants could modify the IFT140 phenotype. Importantly, our studies link a ciliary structural protein to the ADPKD spectrum.

Revisiting the gene mutations and protein profile of WT 9-12: An autosomal dominant polycystic kidney disease cell line.

WT 9-12 is one of the cell lines commonly used for autosomal dominant polycystic kidney disease (ADPKD) studies. Previous studies had described the PKD gene mutations and polycystin expression in WT 9-12. Nonetheless, the mutations occurring in other ADPKD-associated genes have not been investigated. This study aims to revisit these mutations and protein profile of WT 9-12. Whole genome sequencing verified the presence of truncation mutation at amino acid 2556 (Q2556X) in PKD1 gene of WT 9-12. Besides, those variations with high impacts included single nucleotide polymorphisms (rs8054182, rs117006360, and rs12925771) and insertions and deletions (InDels) (rs145602984 and rs55980345) in PKD1L2; InDel (rs1296698195) in PKD1L3; and copy number variations in GANAB. Protein profiles generated from the total proteins of WT 9-12 and HK-2 cells were compared using isobaric tags for relative and absolute quantitation (iTRAQ) analysis. Polycystin-1 was absent in WT 9-12. The gene ontology enrichment and reactome pathway analyses revealed that the upregulated and downregulated proteins of WT 9-12 relative to HK-2 cell line leaded to signaling pathways related to immune response and amino acid metabolism, respectively. The ADPKD-related mutations and signaling pathways associated with differentially expressed proteins in WT 9-12 may help researchers in cell line selection for their studies.

Cleavage fragments of the C-terminal tail of polycystin-1 are regulated by oxidative stress and induce mitochondrial dysfunction.

Mutations in the gene encoding polycystin-1 (PC1) are the most common cause of autosomal dominant polycystic kidney disease (ADPKD). Cysts in ADPKD exhibit a Warburg-like metabolism characterized by dysfunctional mitochondria and aerobic glycolysis. PC1 is an integral membrane protein with a large extracellular domain, a short C-terminal cytoplasmic tail and shares structural and functional similarities with G protein-coupled receptors. Its exact function remains unclear. The C-terminal cytoplasmic tail of PC1 undergoes proteolytic cleavage, generating soluble fragments that are overexpressed in ADPKD kidneys. The regulation, localization, and function of these fragments is poorly understood. Here, we show that a ∼30 kDa cleavage fragment (PC1-p30), comprising the entire C-terminal tail, undergoes rapid proteasomal degradation by a mechanism involving the von Hippel-Lindau tumor suppressor protein. PC1-p30 is stabilized by reactive oxygen species, and the subcellular localization is regulated by reactive oxygen species in a dose-dependent manner. We found that a second, ∼15 kDa fragment (PC1-p15), is generated by caspase cleavage at a conserved site (Asp-4195) on the PC1 C-terminal tail. PC1-p15 is not subject to degradation and constitutively localizes to the mitochondrial matrix. Both cleavage fragments induce mitochondrial fragmentation, and PC1-p15 expression causes impaired fatty acid oxidation and increased lactate production, indicative of a Warburg-like phenotype. Endogenous PC1 tail fragments accumulate in renal cyst-lining cells in a mouse model of PKD. Collectively, these results identify novel mechanisms regarding the regulation and function of PC1 and suggest that C-terminal PC1 fragments may be involved in the mitochondrial and metabolic abnormalities observed in ADPKD.

DNA methyltransferase 1 (DNMT1) promotes cyst growth and epigenetic age acceleration in autosomal dominant polycystic kidney disease.

Alteration of DNA methylation leads to diverse diseases, and the dynamic changes of DNA methylation (DNAm) on sets of CpG dinucleotides in mammalian genomes are termed "DNAm age" and "epigenetic clocks" that can predict chronological age. However, whether and how dysregulation of DNA methylation promotes cyst progression and epigenetic age acceleration in autosomal dominant polycystic kidney disease (ADPKD) remains elusive. Here, we show that DNA methyltransferase 1 (DNMT1) is upregulated in cystic kidney epithelial cells and tissues and that knockout of Dnmt1 and targeting DNMT1 with hydralazine, a safe demethylating agent, delays cyst growth in Pkd1 mutant kidneys and extends life span of Pkd1 conditional knockout mice. With methyl-CpG binding domain (MBD) protein-enriched genome sequencing (MBD-seq), DNMT1 chromatin immunoprecipitation (ChIP)-sequencing and RNA-sequencing analysis, we identified two novel DNMT1 targets, PTPRM and PTPN22 (members of the protein tyrosine phosphatase family). PTPRM and PTPN22 function as mediators of DNMT1 and the phosphorylation and activation of PKD-associated signaling pathways, including ERK, mTOR and STAT3. With whole-genome bisulfide sequencing in kidneys of patients with ADPKD versus normal individuals, we found that the methylation of epigenetic clock-associated genes was dysregulated, supporting that epigenetic age is accelerated in the kidneys of patients with ADPKD. Furthermore, five epigenetic clock-associated genes, including Hsd17b14, Itpkb, Mbnl1, Rassf5 and Plk2, were identified. Thus, the diverse biological roles of these five genes suggest that their methylation status may not only predict epigenetic age acceleration but also contribute to disease progression in ADPKD.

Modulating inflammation with interleukin 37 treatment ameliorates murine Autosomal Dominant Polycystic Kidney Disease.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a leading cause of kidney failure and is associated with substantial morbidity and mortality. Interstitial inflammation is attributed to the action of infiltrating macrophages and is a feature thought to aggravate disease progression. Here, we investigated the therapeutic potential of the anti-inflammatory IL37b cytokine as a treatment for ADPKD using genetic mouse models, demonstrating that transgenic expression of human IL37b reduced collecting duct cyst burden in both early and adult-onset ADPKD rodent models. Moreover, injection of recombinant human IL37b could also reduce cyst burden in early onset ADPKD mice, an observation not associated with increased macrophage number at early stages of cyst formation. Interestingly, transgenic IL37b expression also did not alter macrophage numbers in advanced disease. Whole kidney RNA-seq highlighted an IL37b-mediated upregulation of the interferon signaling pathway and single-cell RNA-seq established that these changes originate at least partly from kidney resident macrophages. We further found that blocking type I interferon signaling in mice expressing IL37b resulted in increased cyst number, confirming this as an important pathway by which IL37b exerts its beneficial effects. Thus, our studies show that IL37b promotes interferon signaling in kidney resident macrophages which suppresses cyst initiation, identifying this protein as a potential therapy for ADPKD.

Molecular genetic diagnosis of kidney ciliopathies: Lessons from interpreting genomic sequencing data and the requirement for accurate phenotypic data.

INTRODUCTION: Massively parallel sequencing (MPS) techniques have made a major impact on the identification of the genetic basis of inherited kidney diseases such as the ciliopathy autosomal dominant polycystic kidney disease (ADPKD). Great care must be taken when analysing MPS data in isolation from accurate phenotypic information, as this can cause misdiagnosis. METHODS: Here, we describe a family trio, recruited to the Genomics England 100,000 Genomes Project, labelled as having cystic kidney disease, who were genetically unsolved following routine data analysis pipelines. We performed a bespoke reanalysis of Whole Genome Sequencing (WGS) data and coupled this with revised phenotypic data and targeted PCR and Sanger sequencing to provide a precise molecular genetic diagnosis. RESULTS: We detected a heterozygous PKD1 frameshift variant within the WGS data which segregated with the redefined ADPKD phenotypes. An additional heterozygous exon deletion in ALG8 was also found in affected and unaffected individuals, but its precise clinical significance remains unclear. CONCLUSION: This case illustrates that reanalysis of WGS data in unsolved cases of cystic kidney disease is valuable. Clinical phenotypes must be reassessed as these may have been incorrectly recorded and evolve over time. Undertaking additional studies including genotype-phenotype correlation in wider family members provides useful diagnostic information.

Pkd2, mutations linking to autosomal dominant polycystic kidney disease, localizes to the endoplasmic reticulum and regulates calcium signaling in fission yeast.

Autosomal dominant polycystic kidney disease (ADPKD) is a renal disorder caused by mutations in the PKD2 gene, which encodes polycystin-2/Pkd2, a transient receptor potential channel. The precise role of Pkd2 in cyst formation remains unclear. The fission yeast Schizosaccharomyces pombe has a putative transient receptor potential channel, Pkd2, which shares similarities with human Pkd2. In this study, truncation analyses of fission yeast Pkd2 were conducted to investigate its localization and function. The results revealed that Pkd2 localizes not only to the plasma membrane but also to the endoplasmic reticulum (ER) in fission yeast. Furthermore, Pkd2 regulates calcium signaling in fission yeast, with the transmembrane domains of Pkd2 being sufficient for these processes. Specifically, the C-terminal region of Pkd2 plays a crucial role in the regulation of calcium signaling. Interestingly, human Pkd2 also localized to the ER and had some impact on calcium signaling in fission yeast. However, human Pkd2 failed to suppress the loss of fission yeast Pkd2. These findings indicate that hPkd2 may not completely substitute for cellular physiology of fission yeast Pkd2. This study provides insights into the localization and functional characteristics of Pkd2 in fission yeast, contributing to our understanding of the pathogenesis of ADPKD.

Visceral Adiposity and Progression of ADPKD: A Cohort Study of Patients From the TEMPO 3:4 Trial.

RATIONALE & OBJECTIVE: Body mass index (BMI) is an independent predictor of kidney disease progression in individuals with autosomal dominant polycystic kidney disease (ADPKD). Adipocytes do not simply act as a fat reservoir but are active endocrine organs. We hypothesized that greater visceral abdominal adiposity would associate with more rapid kidney growth in ADPKD and influence the efficacy of tolvaptan. STUDY DESIGN: A retrospective cohort study. SETTING & PARTICIPANTS: 1,053 patients enrolled in the TEMPO 3:4 tolvaptan trial with ADPKD and at high risk of rapid disease progression. PREDICTOR: Estimates of visceral adiposity extracted from coronal plane magnetic resonance imaging (MRI) scans using deep learning. OUTCOME: Annual change in total kidney volume (TKV) and effect of tolvaptan on kidney growth. ANALYTICAL APPROACH: Multinomial logistic regression and linear mixed models. RESULTS: In fully adjusted models, the highest tertile of visceral adiposity was associated with greater odds of annual change in TKV of≥7% versus<5% (odds ratio [OR], 4.78 [95% CI, 3.03-7.47]). The association was stronger in women than men (interaction P<0.01). In linear mixed models with an outcome of percent change in TKV per year, tolvaptan efficacy (% change in TKV) was reduced with higher visceral adiposity (3-way interaction of treatment ∗ time ∗ visceral adiposity, P=0.002). Visceral adiposity significantly improved classification performance of predicting rapid annual percent change in TKV for individuals with a normal BMI (DeLong's test z score: -2.03; P=0.04). Greater visceral adiposity was not associated with estimated glomerular filtration rate (eGFR) slope in the overall cohort; however, visceral adiposity was associated with more rapid decline in eGFR slope (below the median) in women (fully adjusted OR, 1.06 [95% CI, 1.01-1.11] per 10 unit increase in visceral adiposity) but not men (OR, 0.98 [95% CI, 0.95-1.02]). LIMITATIONS: Retrospective; rapid progressors; computational demand of deep learning. CONCLUSIONS: Visceral adiposity that can be quantified by MRI in the coronal plane using a deep learning segmentation model independently associates with more rapid kidney growth and improves classification of rapid progression in individuals with a normal BMI. Tolvaptan efficacy decreases with increasing visceral adiposity. PLAIN-LANGUAGE SUMMARY: We analyzed images from a previous study with the drug tolvaptan conducted in patients with autosomal dominant polycystic kidney disease (ADPKD) to measure the amount of fat tissue surrounding the kidneys (visceral fat). We had previously shown body mass index can predict kidney growth in this population; now we determined whether visceral fat was an important factor associated with kidney growth. Using a machine learning tool to automate measurement of fat in images, we observed that visceral fat was independently associated with kidney growth, that it was a better predictor of faster kidney growth in lean patients than body mass index, and that having more visceral fat made treatment of ADPKD with tolvaptan less effective.

A Case of Autosomal Dominant Polycystic Kidney Disease With Resolution of Massive Pericardial Effusion After Renal Transcatheter Artery Embolization.

A 68-year-old woman being treated with hemodialysis for autosomal dominant polycystic kidney disease was admitted for progressive dyspnea over 6 months. On chest radiography, her cardiothoracic ratio had increased from 52.2% 6 months prior, to 71%, and echocardiography revealed diffuse pericardial effusion and right ventricular diastolic insufficiency. A resultant pericardial tamponade was thought to be the cause of the patient's dyspnea, and therefore a pericardiocentesis was performed, with a total of 2,000mL of fluid removed. However, 21 days later the same amount of pericardial fluid had reaccumulated. The second pericardiocentesis was performed, followed by transcatheter renal artery embolization (TAE). The kidneys, which were hard on palpation before TAE, softened immediately after TAE. After resolution of the pericardial effusion was confirmed, the patient was discharged after 24 days in hospital. Twelve months later, the patient was asymptomatic, the cardiothoracic ratio decreased to 48% on chest radiography and computed tomography revealed no reaccumulation of pericardial effusion. This case illustrates a potential relationship between enlarged kidneys in autosomal dominant polycystic kidney disease and pericardial effusion.

Serum bile acids associate with liver volume in polycystic liver disease and decrease upon treatment with lanreotide.

BACKGROUND: Polycystic liver disease (PLD) is a common extrarenal manifestation of autosomal dominant polycystic kidney disease (ADPKD). Bile acids may play a role in PLD pathogenesis. We performed a post-hoc exploratory analysis of bile acids in ADPKD patients, who had participated in a trial on the effect of a somatostatin analogue. Our hypothesis was that serum bile acid levels increase in PLD, and that lanreotide, which reduces liver growth, may also reduce bile acid levels. Furthermore, in PLD, urinary excretion of bile acids might contribute to renal disease. METHODS: With liquid chromatography-mass spectrometry, 11 bile acids in serum and 6 in urine were quantified in 105 PLD ADPKD patients and 52 age-, sex-, mutation- and eGFR-matched non-PLD ADPKD patients. Sampling was done at baseline and after 120 weeks of either lanreotide or standard care. RESULTS: Baseline serum levels of taurine- and glycine-conjugated bile acids were higher in patients with larger livers. In PLD patients, multiple bile acids decreased upon treatment with lanreotide but remained stable in untreated subjects. Changes over time did not correlate with changes in liver volume. Urine bile acid levels did not change and did not correlate with renal disease progression. CONCLUSION: In ADPKD patients with PLD, baseline serum bile acids were associated with liver volume. Lanreotide reduced bile acid levels and has previously been shown to reduce liver volume. However, in this study, the decrease in bile acids was not associated with the change in liver volume.

Fluid shear stress stimulates ATP release without regulating purinergic gene expression in the renal inner medullary collecting duct.

In the kidney, the flow rate of the pro-urine through the renal tubules is highly variable. The tubular epithelial cells sense these variations in pro-urinary flow rate in order to regulate various physiological processes, including electrolyte reabsorption. One of the mechanosensitive pathways activated by flow is the release of ATP, which can then act as a autocrine or paracrine factor. Increased ATP release is observed in various kidney diseases, among others autosomal dominant polycystic kidney disease (ADPKD). However, the mechanisms underlying flow-induced ATP release in the collecting duct, especially in the inner medullary collecting duct, remain understudied. Using inner medullary collecting duct 3 (IMCD3) cells in a microfluidic setup, we show here that administration of a high flow rate for 1 min results in an increased ATP release compared to a lower flow rate. Although the ATP release channel pannexin-1 contributed to flow-induced ATP release in Pkd1-/- IMCD3 cells, it did not in wildtype IMCD3 cells. In addition, flow application increased the expression of the putative ATP release channel connexin-30.3 (CX30.3) in wildtype and Pkd1-/- IMCD3 cells. However, CX30.3 knockout IMCD3 cells exhibited a similar flow-induced ATP release as wildtype IMCD3 cells, suggesting that CX30.3 does not drive flow-induced ATP release in wildtype IMDC3 cells. Collectively, our results show differential mechanisms underlying flow-induced ATP release in wildtype and Pkd1-/- IMCD3 cells and further strengthen the link between ADPKD and pannexin-1-dependent ATP release.

Atypical noncontiguous TSC2/PKD1 gene deletions presenting as tuberous sclerosis/polycystic kidney disease contiguous gene syndrome.

Tuberous sclerosis complex (TSC) and autosomal dominant polycystic kidney disease (ADPKD) are genetically distinct disorders typically associated with pathogenic variants in TSC1 and TSC2 for the former and PKD1 and PKD2 for the latter. TSC2 and PKD1 lie adjacent to each other, and large deletions comprising both genes lead to TSC2/PKD1 contiguous gene deletion syndrome (CGS). In this study, we describe a young female patient exhibiting symptoms of TSC2/PKD1 CGS in which genetic analysis disclosed two noncontiguous partial gene deletions in TSC2 and PKD1 that putatively are responsible for the manifestations of the syndrome. Further analysis revealed that both deletions appear to be de novo on the maternal chromosome, presumably with a germline origin. Despite extensive analysis, no maternal chromosomal rearrangement triggering these pathogenic variants was detected. This case elucidates a unique pathogenesis for TSC2/PKD1 CGS, diverging from the common contiguous deletions typically observed, marking the first reported instance of TSC2/PKD1 CGS caused by independent, functionally significant partial gene deletions.

Effects of salt and protein intake on polyuria in V2RA-treated ADPKD patients.

BACKGROUND: The only treatment proven to be renoprotective in autosomal dominant polycystic kidney disease (ADPKD) is a vasopressin V2-receptor antagonist (V2RA). However, aquaresis-associated side effects limit tolerability. We investigated whether salt and/or protein intake influences urine volume and related endpoints in V2RA-treated ADPKD patients. METHODS: In this randomized, controlled, double-blind, crossover trial, ADPKD patients treated with maximally tolerated dose of a V2RA were included. While on a low salt and low protein diet, patients were given additional salt and protein to mimic regular intake, which was subsequently replaced by placebo in random order during four 2-week periods. Primary endpoint was change in 24-h urine volume. Secondary endpoints were change in quality of life, measured glomerular filtration rate (mGFR), blood pressure and copeptin level. RESULTS: Twelve patients (49 ± 8 years, 25.0% male) were included. Baseline salt and protein intake were 10.8 ± 1.3 g/24-h and 1.2 ± 0.2 g/kg/24-h, respectively. During the low salt and low protein treatment periods, intake decreased to 5.8 ± 1.6 g/24-h and 0.8 ± 0.1 g/kg/24-h, respectively. Baseline 24-h urine volume (5.9 ± 1.2 L) decreased to 5.2 ± 1.1 L (-11%, P = .004) on low salt and low protein, and to 5.4 ± 0.9 L (-8%, P = .04) on low salt. Reduction in 24-h urine volume was two times greater in patients with lower urine osmolality (-16% vs -7%). Polyuria quality of life scores improved in concordance with changes in urine volume. mGFR decreased during the low salt and low protein, while mean arterial pressure did not change during study periods. Plasma copeptin decreased significantly during low salt and low protein periods. CONCLUSION: Lowering dietary salt and protein intake has a minor effect on urine volume in V2RA-treated ADPKD patients. Reduced intake of osmoles decreased copeptin concentrations and might thus increase the renoprotective effect of a V2RA in ADPKD patients.

Combining genotype with height-adjusted kidney length predicts rapid progression of ADPKD.

INTRODUCTION: Our main objective was to identify baseline prognostic factors predictive of rapid disease progression in a large unselected clinical autosomal dominant polycystic kidney disease (ADPKD) cohort. METHODS: A cross-sectional analysis was performed in 618 consecutive ADPKD patients assessed and followed-up for over a decade. A total of 123 patients (19.9%) had reached kidney failure by the study date. Data were available for the following: baseline eGFR (n = 501), genotype (n = 549), baseline ultrasound mean kidney length (MKL, n = 424) and height-adjusted baseline MKL (HtMKL, n = 377). Rapid disease progression was defined as an annualized eGFR decline (∆eGFR) of >2.5 mL/min/year by linear regression over 5 years (n = 158). Patients were further divided into slow, rapid and very rapid ∆eGFR classes for analysis. Genotyped patients were classified into several categories: PKD1 (T, truncating; or NT, non-truncating), PKD2, other genes (non-PKD1 or -PKD2), no mutation detected or variants of uncertain significance. RESULTS: A PKD1-T genotype had the strongest influence on the probability of reduced baseline kidney function by age. A multivariate logistic regression model identified PKD1-T genotype and HtMKL (>9.5 cm/m) as independent predictors for rapid disease progression. The combination of both factors increased the positive predictive value for rapid disease progression over age 40 years and of reaching kidney failure by age 60 years to 100%. Exploratory analysis in a subgroup with available total kidney volumes showed higher positive predictive value (100% vs 80%) and negative predictive value (42% vs 33%) in predicting rapid disease progression compared with the Mayo Imaging Classification (1C-E). CONCLUSION: Real-world longitudinal data confirm the importance of genotype and kidney length as independent variables determining ∆eGFR. Individuals with the highest risk of rapid disease progression can be positively selected for treatment based on this combination.

The inhibition of MDM2 slows cell proliferation and activates apoptosis in ADPKD cell lines.

INTRODUCTION: Autosomal dominant polycystic kidney disease (ADPKD) is characterised by progressive cysts formation and renal enlargement that in most of cases leads to end stage of renal disease (ESRD). This pathology is caused by mutations of either PKD1 or PKD2 genes that encode for polycystin-1 (PC1) and polycystin-2 (PC2), respectively. These proteins function as receptor-channel complex able to regulate calcium homeostasis. PKD1/2 loss of function impairs different signalling pathways including cAMP and mTOR that are considered therapeutic targets for this disease. In fact, Tolvaptan, a vasopressin-2 antagonist that reduces cAMP levels, is the only drug approved for ADPKD treatment. Nevertheless, some ADPKD patients developed side effects in response to Tolvaptan including liver damage. Conversely, mTOR inhibitors that induced disease regression in ADPKD animal models failed the clinical trials. RESULTS: Here, we show that the inhibition of mTOR causes the activation of autophagy in ADPKD cells that could reduce therapy effectiveness by drug degradation through the autophagic vesicles. Consistently, the combined treatment with rapamycin and chloroquine, an autophagy inhibitor, potentiates the decrease of cell proliferation induced by rapamycin. To overcome the dangerous activation of autophagy by mTOR inhibition, we targeted MDM2 (a downstream effector of mTOR signalling) that is involved in TP53 degradation by using RG7112, a small-molecule MDM2 inhibitor used for the treatment of haematologic malignancies. The inhibition of MDM2 by RG7112 prevents TP53 degradation and increases p21 expression leading to the decrease of cell proliferation and the activation of apoptosis. CONCLUSION: The targeting of MDM2 by RG7112 might represent a new therapeutic option for the treatment of ADPKD.