PKDCON

agenda

Speakers

Posters

FAQs

PKDCON

agenda

Speakers

Posters

FAQs

Poster sessions in 2018 were a great opportunity to talk to the researchers in the PKD space. PKDCON 2022 will look different, but the virtual platform won’t keep us from interacting with researchers. The information below can give you an idea of the great work these researchers are doing. Reading these ahead of the conference is a great way to get ready for some great conversations with researchers!

Once you register, you can view the posters on our virtual event platform. Email research@pkdcure.org if you have any questions! 

poster summaries

Alex Yashchenko

Institution
(1) Division of Nephrology, University of Oklahoma Health Sciences Center
(2) Department of Cell, Developmental, and Integrative Biology, University of Alabama at Birmingham
(3) Singapore Immunology Network (SIgN), Agency for Science, Technology & Research

Co-authors
Sarah Bland (1), Zhang Li (2), Florent Ginhoux (3), Kurt Zimmerman (1)

Unravelling the heterogeneity of kidney resident macrophages using single cell RNA sequencing

Macrophages differ across tissues based on function, origin, and localization. However, recent findings suggest that macrophage subpopulations also exist within tissues and may have distinct roles. We used single cell RNA sequencing to identify several kidney resident macrophage (KRM) subpopulations. We confirmed that one subpopulation, the Ccr2+ KRM, was of recent monocyte origin and localized preferentially to the renal cortex. We also identified several genes that may be required for the differentiation of monocytes into Ccr2+ KRM, and confirmed that loss of one gene, Cx3cr1, resulted in decreased quantity of Ccr2+ KRM. We applied these findings to a mouse model of cystic kidney disease and showed that loss of Cx3cr1 reduced disease progression. These findings suggest that several macrophage subpopulations are present in the kidney and may be uniquely involved in disease progression.
Diana Bruen

Institution
(1) Santa Barbara Nutrients, Inc
(2) RenAlign
(3) Department of Molecular, Cellular, and Developmental Biology and Neuroscience Research Institute, University of California Santa Barbara

Co-authors
Jacob J. Kingaard (1), Meg Munits (1), Clarissa S. Paimanta (2), Jacob A. Torres (1,3), Jessianna Saville (2), Thomas Weimbs (1,3)

Ren.Nu, a dietary program for individuals with autosomal-dominant polycystic kidney disease implementing a sustainable, plant-focused, kidney-safe, ketogenic approach with avoidance of renal stressors

In this study, we examined the feasibility, and overall experience, of ADPKD participants in the Beta Ren.Nu program.  Ren.Nu is a 12-week, dietitian-supervised, virtual program that teaches individuals with ADPKD how to implement a plant-focused ketogenic diet while avoiding kidney stressors that may worsen disease progression. The majority of participants, who completed the program, reported improvements in their health and well-being including pain levels, weight loss, hypertension and eGFR. Adherence to the program was very high and the feasibility of the dietary and lifestyle changes was rated highly. This study indicates that Ren.NU may be a viable nutrition intervention to slow or prevent ADPKD progression and improve health outcomes.

Robin Maser

Institution
University of Kansas Medical Center

Co-authors
Ericka Nevarez Munoz,
Brenda Magenheimer

ADPKD Mutations in the Stalk/Tethered Agonist of Polycystin-1 CTF Affect Signaling and GPS Cleavage

Polycystin-1, the product of the PKD1 gene, is a membrane receptor capable of activating multiple signaling pathways to alter the function/properties of the cell. Polycystin-1 resembles Adhesion GPCRs that undergo an unusual, self-catalyzed cleavage resulting in N-terminal (NTF) and C-terminal (CTF) fragments. We recently showed that the short sequence region at the beginning of the PC1 CTF – the stalk – acts as a tethered peptide ligand which ‘bends backwards’ to bind to the remainder of the CTF, resulting in activation of a signaling reporter. We have now analyzed all of the known mutations/polymorphisms within the stalk region of the PC1 CTF for their effect on signaling and cleavage in order to better understand the tethered ligand mechanism and its potential role in PKD. 

Edmund Chun Yu Lee

Institution

(1) Regulus Therapeutics Inc.
(2) Department of Internal Medicine and Division of Nephrology, University of Texas Southwestern Medical Center
(3) Berman Consulting
(4) PharmaDRS Consulting

Co-authors
Tania Valencia (1), Tate Owen (1), Andrea Flaten (2), Harini Ramalingam (2), Vishal Patel (2), Francesca Varrone (1), Laura Ruff (1), Cindy Berman (3), Rekha Garg (4), Denis Drygin (1)

Discovery and Characterization of the Next-generation Anti-miR-17 Oligonucleotide RGLS8429 for the Treatment of ADPKD

ADPKD is mostly caused by mutations in PKD1 and PKD2 genes, which reduce levels of their encoded proteins polycystin-1 (PC1) and polycystin-2 (PC2), and result in kidney cyst formation. Levels of PC1 and PC2 in urinary exosomes are lower in ADPKD patients than healthy volunteers, and correlate inversely with disease severity [1]. Unfortunately, treatment options for ADPKD are limited and inadequate.

Here we report the discovery and characterization of a new drug for ADPKD called RGLS8429. RGLS8429 is the next-generation anti-miR-17 oligonucleotide designed to inhibit the activity of the miR-17 family of miRNAs. miR-17 is overexpressed in ADPKD kidney cysts, where it further decreases PC1 and PC2 levels [2-4]. In preclinical studies, inhibition of miR-17 increased PC1 and PC2 levels, inhibited kidney cyst growth, and improved kidney function [3-5].

A clinical trial evaluating RGLS8429 in heathy volunteers is anticipated to start in Q2 2022.

Tania Valencia

Institution
Regulus Therapeutics Inc.

Co-authors
Tate Owen (1), Wendi Lea, Christopher Ward, Edmund Chun Yu Lee, Karl Cremer, Claire Padgett, Rekha Garg, Denis Drygin

Results from the Phase 1b Clinical Trial of RGLS4326, the first-generation anti-miR-17 oligonucleotide, and Study Design for the Phase 1b Clinical Trial of RGLS8429, the next-generation anti-miR-17 oligonucleotide, for the Treatment of Patients with ADPKD

ADPKD is caused by mutations in PKD1 or PKD2 genes, which reduces level of their encoded-proteins polycystin-1 (PC1) and polycystin-2 (PC2) in urinary exosomes (uELV) of ADPKD patients compared to healthy volunteers. Importantly,  uELV PC1 and PC2 levels inversely correlates with disease severity.

Here we report results from the Phase 1b trial of RGLS4326, the first-generation anti-miR-17 oligonucleotide, for treatment of ADPKD. In this study, 4 subcutaneous injections of 1 or 0.3 mg/kg of RGLS4326 every other week was well tolerated in ADPKD patients. Importantly, significant increase in uELV PC1 and PC2 levels were observed in patients who received 1 mg/kg of RGLS4326.

The next-generation anti-miR-17 oligonucleotide RGLS8429, specifically designed to eliminate dose-limiting CNS toxicity previously observed in RGLS4326 studies in mice and monkeys, is currently being developed for treatment of ADPKD. Here we discuss study design for the anticipated Phase 1 clinical trial of RGLS8429 in ADPKD patients.

Aurora Daniele (1,3)

Institution
(1) CEINGE-Biotecnologie Avanzate Scarl, Via G. Salvatore 486, 80145, Napoli, Italy;
(2) Dipartimento di Scienze e Tecnologie Ambientali, Biologiche, Farmaceutiche, Università della Campania “Luigi Vanvitelli”, via Vivaldi 43, 81100, Caserta, Italy.
(3) Dipartimento di Sanità Pubblica, Unità di Nefrologia, Università di Napoli ‘Federico II’, Napoli, Italy

Co-authors
Daniela D’Arco (1), Marta Mallardo (1,2), Ersilia Nigro (1,2), Maria Amicone (3),
Eleonora Riccio (3), Antonio Pisani (3)

Next Generation Sequencing for the diagnosis of polycystic kidney disease in an Italian cohort of patients

The clinical diagnosis of PKD is established by family history and renal imaging modalities. However, these diagnostic tests are often ambiguous, particularly in young individuals. Consequently, genetic testing plays an increasingly important role in the diagnosis and management of patients with PKD. Moreover, with the development of potentially effective pharmacologic treatments for PKD, the need for accurate diagnostic genetic tests has become more compelling.  On the genetic point of view, PKD can be inherited as an autosomal dominant trait (ADPKD) that is mainly caused by mutations in two large genes, PKD1 and PKD2, accounting for about 75% and 25% of cases, respectively, in clinically well-characterized populations. PKD1 spans 46 exons and encodes polycystin-1 with 4303 amino acids. PKD2 spans 15 exons, encoding polycystin-2, encoding 968 amino acids.  However, genetic analysis is complicated by the presence of six PKD1 pseudogenes, by the large gene sizes, and by allelic heterogeneity that share 97.7% sequence identity with the PKD1 gene exons 1 to 33. In addition, there is also an autosomal recessive polycystic kidney disease (ARPKD) that is a fibrocystic hepatorenal disease representing a rare but the most severe PKD form and characterized by an early end stage renal disease and high morbidity and mortality caused by mutations in PKHD1 gene. The aim of our study was to develop and validate a rapid and cost-saving genetic test for the molecular PKD diagnosis through a Next Generation Sequencing (NGS) approach to improve patients’ diagnosis, prognosis, surveillance and therapy by performing differential diagnosis, identifying new causative mutations and clarifying the genotype–phenotype correlations. In this context, we established and validated a NGS panel of 20 genes causing PKD and analyzed a cohort of 120 patients from Nephrology Unit, Department of Public Health, Federico II  University, Naples. Our results revealed mutations in PKD1, PKD2, and PKHD1 genes in 56%, 15.5%, 1.4% of affected patients respectively, with a detection rate of 83%. We found the following type of mutations: nonsense, missense, frameshift, stop loss, splicing, in frame with the prevalence of missense and nonsense mutations. PKD1 resulted the most mutated gene with 70% of total mutations, distributed in all exons, without a mutational hot spot. However, taken together, exons 15, 5 and 42 accounted for 45% of total mutations in PKD1. In summary, NGS-based PKD genetic analysis is a highly accurate and reliable approach for mutation analysis, achieving high sensitivity and improved intronic coverage with a faster turnaround time and lower cost. Optimization of the workflow and the stepwise process quality control metrics for data analysis will likely become routine for clinical genetic testing, and NGS would be an appropriate standard for clinical genetic testing of PKD. These results allow the identification and classification of genetic defects that have an important impact on the management and genetic counselling of the patients. 

Rudi Vennekens

Institution
Flemish Institute of Biotechnology

Properties of sensory neurons in ADPKD, and their contribution to ADPKD related chronic pain

A very common symptom and patient reported outcome in ADPKD is acute and chronic pain. A recent study with patients and clinicians in the United States, Europe and Japan found that ADPKD-related pain was the most important outcome impacting physical functioning, with complex and distinctive presentations ranging from feeling full/discomfort to acute sharp pain. However, the source of ADPKD-related pain is poorly understood, especially for chronic pain. Therefore, pain in ADPKD patients is often under-recognized and inadequately managed. Yet, chronic, long-term pain is known to affect at least 2 in 3 ADPKD patients and is associated with reduced quality of life, often intense distress, and considerable economic and healthcare costs from emergency hospital admissions, polypharmacy (antiobiotics and pain relief including morphine) and impact on work/home life.

No gold-standard modality for pain relief exists in patients with ADPKD. Indeed, the current pharmacological analgesia management in ADPKD is prescribed according to the WHO three-step analgesic ladder. However, an important caveat in this ADPKD population is the nephrotoxicity of NSAID’s and the impaired clearance of opioids, limiting their use and safety. Patients with unsatisfactorily pain relief will undergo more invasive procedures ranging from cyst aspiration, renal denervation to radical nephrectomy.

Despite the scale and burden, there has been little research into ADPKD associated pain and its treatment. Why pain occurs is inadequately understood – it is not always related to kidney size and may occur in adolescence and early stages when no other symptoms are present.

One attractive strategy is to intercept pain at the source, by targeting the sensory neurons that generate nociceptive signals. Currently, there is virtually no knowledge on the properties of sensory neurons or molecular sensory receptors in the context of ADPKD. Considering that PKD1 and PKD2 (the two predominant genes in which mutations cause ADPKD) are expressed in sensory neurons, that mutations in PKD1 and PKD2 lead to abnormalities in cellular signaling in various cell types, and that secretion of a variety of factors (such as MCP1) which influence sensory neurons is altered in ADPKD, we hypothesize that the properties of nociceptive sensory neurons are distorted in the context of ADPKD, which might contribute to the precipitation of chronic pain in ADPKD patients and could even contribute to development of the disease.

Michael Caplan

Institution
Yale University

Can a fragment of polycystin-1 unlock the door to gene therapy for ADPKD?

ADPKD is the most prevalent potentially lethal monogenic disorder. Approximately 78% of cases are caused by mutations in the PKD1 gene, which encodes polycystin-1 (PC1). PC1 is a large 462-kDa protein that undergoes cleavage in its N and C-terminal domains. C-terminal cleavage produces fragments that translocate to mitochondria. We have found that transgenic expression of a protein corresponding to the final 200 amino acid residues of PC1 in a Pkd1-knockout orthologous murine model of ADPKD dramatically suppresses the cystic phenotype and preserves renal function. This suppression depends upon an interaction between the C-terminal tail of PC1 and the mitochondrial enzyme Nicotinamide Nucleotide Transhydrogenase. Expression of the PC1-CTT stimulates NNT activity.

Recent data from Dr. S. Somlo’s laboratory demonstrate that re-expression of full length PC1 or PC2 can dramatically reverse advanced cystic disease attributable to inactivation of Pkd1 or Pkd2, respectively. These data suggest the intriguing possibility that development of an intervention that produces expression of functional PC proteins in the renal epithelial cells of patients could potentially not only prevent disease progression but could also diminish the severity of established cystic pathology. The development and testing of such an intervention are complicated by the fact that the cDNA encoding full length PC1 is far too large to be packaged in any currently available conventional viral gene delivery vector. Our data show that initiation of conditional PC1-CTT expression simultaneously with conditional disruption of Pkd1 substantially dampens the development of the ADPKD phenotype in a mouse model. In Aim 1 we will determine whether expression of the PC1-CTT recapitulates the capacity of the full length PC1 protein to induce the resolution of extant cysts. Were this to be the case it would constitute a strong proof of concept for the development of gene therapy strategies built around the delivery of the sequence encoding the PC1-CTT, since this sequence is only ~600 bp in length.

In Aim 2 we will take advantage of in vitro assays to define the minimal active piece of the PC1-CTT that is needed in order to mediate its effects on NNT activity and on cyst formation. The sequence that encodes the PC1-CTT and the sequence that encodes its minimal active piece will be employed in the creation of viral gene delivery vectors. We will test the capacity of these vectors to drive the expression of the encoded proteins in the epithelial cells of mouse kidneys in vivo. Will also assess whether administration of these vectors is able to slow or reverse cystic disease in orthologous mouse models of ADKPD. A demonstration that delivery of the PC1-CTT or its minimal active piece reduces ADPKD severity in vivo would provide strong support for further translational development of this approach.

Alison Kriegel

Institution
The Medical College of Wisconsin

Targeting Tolvaptan-Resistant Mechanisms of Fibrosis in ADPKD

ADPKD is a systemic disorder in which kidneys become large due to tubular cyst expansion. This is accompanied by fibrosis (scarring) in the kidney. Tolvaptan is the only FDA-approved drug for treatment of ADPKD. It targets the V2 vasopressin receptor, causing the kidney to reabsorb less water from the urine. While tolvaptan reduces cyst size, modestly slowing the rate of declining renal function, the NIH sponsored “CRISP” study found that the decline in renal function in people with ADPKD correlated with the development of renal fibrosis, not renal size. There are no approved therapies – nor late-stage clinical candidates in development – to address fibrosis in ADPKD. PKD-associated pathology leads to alterations in blood flow and changes in the distribution of cell types within the kidney, including an invasion of inflammatory cells and proliferation of the interstitial cells. It has been proposed that rapid cyst expansion compresses blood flow, contributing to low oxygen in tissues and fibrosis. Our preliminary data our mcwPkd1(nl/nl) model of ADPKD shows that tolvaptan reduces cyst expansion, but not fibrosis. This suggests that fibrosis will be a persistent problem in people with ADPKD, even with tolvaptan treatment. Importantly, this indicates that specific therapies targeting this fibrotic mechanism, rather than cyst formation and growth alone, must be developed to affect progression of renal disease and loss of function in ADPKD. The mcwPkd1(nl/nl) provides an experimental model to dissociate and study causes of fibrosis that are independent of cyst expansion.
The “central dogma” of biology has been that that DNA is transcribed to RNA, which is translated into a functional protein. Some RNAs, called non-coding RNAs, have other functions. MicroRNAs (miRs) are very short, non-coding RNAs regulated like many protein coding genes. They are understood to bind and inhibit RNA translation. A single miR can targeting multiple RNAs causing broad regulatory effects. Our preliminary data indicate that therapeutic targeting of fibrotic signals, in addition to cyst expansion, is needed to slow progression of renal disease and loss of function in ADPKD. This proposal will focus on the role of transforming growth factor beta-1 (TGFB1) and miR-382, a miR increased in by TGFB1, on persistent fibrosis in conjunction with tolvaptan treatment in the mcwPkd1(nl/nl) model of ADPKD. Both are pro-fibrotic signaling factors, that are increased during periods of rapid fibrotic expansion in Pkd1(nl/nl) mouse models. We anticipate these studies will yield important information about the mechanisms regulating fibrosis. Further, this project has the potential to solidify the involvement of specific signaling pathways in the development of renal fibrosis. This knowledge could help develop new, focused therapeutic strategies to slow or preventing morbidity resulting from progressive fibrotic remodeling in people living with ADPKD.

Madhulika Sharma

Institution
The University of Kansas Medical Center

The role of ferritin in polycystic kidney disease

Autosomal Dominant Polycystic Kidney disease (ADPKD) is a genetic disease which is caused by mutations in PKD1, PKD2 or sometimes other genes. The disease manifests with the formation of huge cysts in the kidney. Initially, the disease progresses slowly and can go unnoticed until a person is 25 to 30 years old when symptoms such as elevated blood pressure, flank pain and increase in kidney volume can be noticed. The disease progression depends on the kind of mutation, each with varying level of severity. Regardless of the mutation, there are some common molecular pathways that are affected. The discovery of these pathways is important because manipulations in these pathways may serve as a strategy to treat the disease. We have recently discovered that ferritin, a protein which maintains iron homeostasis is abnormally expressed in the collecting duct cells (CD) (kidney segment where cysts are present) and inflammatory cells (macrophages) of ADPKD kidneys. To determine the role of ferritin, we cultured cyst lining cells from ADPKD patients in a 3D collagen gel where they formed cysts. Ferritin treatment on these cysts resulted in enlargement of cysts. Cell culture studies further revealed that cyst lining cells have increased capacity for ferritin uptake which induces inflammatory pathway (NF-kappa B) activation. These results suggest that ferritin aids in cyst growth. In the first aim of this proposal, we will find if NF-kappa B activation via ferritin induces cell proliferation and inflammation. We will study the effect of ferritin treatment on disease progression in a mouse model of ADPKD. In the second aim, we will delete ferritin from the CD of ADPKD mice and determine if disease progression slows down. We will also delete ferritin from macrophages of these mice to study if these cells contribute to disease progression. If our goals are successful, the study will provide a basis for treating the disease with ferritin inhibitors.

Katherine Dell (1,2)

Institution
(1) Case Western Reserve University School of Medicine
(2) Center for Pediatric Nephrology and Hypertension, Cleveland Clinic Children’s

Co-authors
Chris Flask (1), Elise Keshock (1), Susan Farr (1), Christina MacAskill (1), Jenna Hach (2)

Novel Magnetic Resonance Fingerprinting of Congenital Hepatic Fibrosis (CHF) in ARPKD Patients

ARPKD affects approximately 1/20,000 children and has two main features, polycystic kidneys and the liver disease, congenital hepatic fibrosis (CHF). CHF results in progressive deleterious changes in the bile ducts of the liver and can be associated with significant, life-threatening complications, including portal hypertension (leading to severe bleeding), bile duct infection or cancer. Although kidney disease is common in many ARPKD patients early in life, CHF may not be evident until later in childhood or adulthood. As more ARPKD patients survive after kidney transplantation, significant CHF is becoming more common. Unfortunately, there are currently no disease-specific therapies and treatment is focused on addressing CHF complications. Several novel therapies have shown efficacy in animal models, but have not been studied in ARPKD patients due to the absence of safe and reliable measures of CHF progression. Newer ultrasound (US)-based elastography methods, that measure liver stiffness (scarring) can distinguish mild vs severe CHF but may lack sensitivity to detect and measure early CHF, when therapies are most likely to be effective. Our collaborative research team (Drs. Dell & Flask) has been studying novel MRI methods to assess ARPKD kidney and liver disease progression for over a decade. In previous studies in an ARPKD animal model, we showed that T1 mapping is a sensitive measure of progressive CHF. While encouraging, one major limitation to MRI is that it usually requires long scan times and is affected by movement, which necessitates sedation/anesthesia for many children. This increases risk and would likely prevent participation of younger ARPKD children in clinical trials that use MRI. To address this important limitation, our group has applied and optimized a novel technique, MR-Fingerprinting (MRF), to study both ARPKD kidney and liver disease. MRF allow for rapid and simultaneous acquisition of multiple imaging parameters, including T1 and T2, and is resistant to motion artifact. In a current NIH R01 longitudinal kidney imaging study, we obtained kidney MRF results in ARPKD patients with excellent repeatability and no need for intravenous contrast or sedation. With supplemental funding, we obtained initial liver MRF images, showing that mean T1 values are significantly higher in ARPKD patients with advanced CHF vs. both healthy volunteers and ARPKD patients with milder disease. The overall goal of this project is to establish liver T1-MRF as a safe, sensitive and reproducible CHF imaging biomarker that would facilitate design and implementation of future clinical trials. The Specific Aims are to evaluate T1-MRF across a spectrum of ARPKD liver disease and compare T1-MRF with US measures of liver scarring. The proposed studies would provide a key element needed to conduct clinical trials of CHF treatments and ultimately improve outcomes for ARPKD patients.

Zhang Li

Institution
University of Alabama at Birmingham

Cisplatin-induced renal injury promotes cyst formation in both cilia mutant and Pkd2 mutant mouse models

Introduction: Links between cilia dysfunction, cyst formation, and renal injuries have been reported. In animal models, injury (e.g. ischemia reperfusion injury (IRI) exacerbated the rate of cyst formation suggesting one function of the cilium is to regulate injury response and repair processes. Cisplatin is an antitumor drug used widely in the treatment of varieties of malignancies that also has severe nephrotoxicity side effects. Here we evaluate whether a second form of renal injury induced by cisplatin treatment also leads to mal-repair of the kidney and to increased cyst formation in mouse models with cilia function perturbation. 

Methods: To test the effects of cisplatin-induced renal injury on cyst formation, we utilized a single-dose cisplatin administration protocol (one 9.0mg/kg body weight  injection on adult-induced conditional(CAG-CREERT2) Ift88 (cilia mutant), Pkd2 (cilia dysfunction) and Ift88;Pkd2 double mutant mouse models. To evaluate the impact of cilia disruption on renal injury and repair induced by cisplatin, and its consequences on cyst formation, we preformed immunofluorescence staining for the injury marker SOX9  from wild type and cilia dysfunctional kidneys 14 days or 28 days after cisplatin treatment. Cystic phenotypes were analyzed 8 weeks after cisplatin treatment in Ift88, Pkd2 and Ift88;Pkd2 double mutant mice to evaluate the consequence of cisplatin-induced injury on cyst formation.

Results: Levels of intrinsic/baseline injury were evaluated by analyzing the number of Sox9+ cells. Under baseline conditions, there was a marked increase in the number of Sox9+ cells in the PKD2 mutant kidneys.  The number of Sox9+ cells in the Ift88 or double mutants was slight increased compared to controls, but not nearly to the extent observed in the Pkd2 mutants.  At 14 days post cisplatin injury, the number of Sox9+ cells increased in control and Pkd2 kidneys compared to the basal state, and both were to similar levels.  In contrast, in the Ift88 and double mutants the injury induced increase in Sox9+ cells were blunted compared to what was observed in the Pkd2 mutants and controls. Our preliminary analysis of Pkd2 mutants at 28 days post injury indicated that the number of Sox9+ cells remain elevated compared to controls, suggesting that in Pkd2 mutants, the cells are not undergoing normal repair.  The analysis of the Ift88 and double mutants at 28 days is still in process.  Histological analysis indicate there was a significant increase in cyst severity in all three lines of mutant kidneys at 8 weeks after cisplatin treatment comparted to vehicle treated group. The cyst severity in Pkd2 mutants after cisplatin was markedly enhanced compared to those in Ift88 or double mutant kidneys. 

Conclusions: These data suggest that loss of Pkd2 results in an increased level of injury at baseline and a persistent injury following cisplatin administration (at least to 28 days). This response to injury is reduced by loss of cilia in either Ift88 or double mutants. Finally, comparison of the injury response to the severity of the cysts that form in each model indicate a direct correlation. These data raise the possibility that PKD2 functions to repress an injury response mediated through the cilium and that the cysts that form in these models may be derived from cells that are not able to correctly repair after injury.  

Jonathon Walsh

Institution
(1) Rutgers University
(2) Worcester Polytechnic Institute

Co-authors
Juan Wang (1), Inna Nikonorova (1), Molly DeHart (1), Jagan Srinivasan (2), Maureen Barr (1)

RC- but not N-terminal proteolytic product of polycystin-1 colocalizes with polycystin-2 on cilia and ciliary EVs in C. elegans

Mutations in polycystin-1 and polycystin-2 result in autosomal dominant polycystic kidney disease (ADPKD), one of the most common ciliopathies and the most common cause of adult-onset renal failure. Cilia are cellular organelles for both receiving signals and broadcasting signals, the latter via ciliary extracellular vesicles (EVs). EVs are tiny membrane-bound vesicles that deliver lipids, nucleic acids, and proteins, and are information carriers of central importance for intercellular signaling. Polycystin-1 and polycystin-2 localize to cilia and EVs in an evolutionarily conserved manner, hinting at a biological importance of polycystin-carrying EVs. Polycystin-1 undergoes autoproteolysis at the GPCR proteolysis site (GPS) in the juxta of the large N-terminal extracellular domain and the 11 transmembrane domains. Mutations in the GPS are pathogenic, however, the physiological function of polycystin-1 proteolytic fragments in cilia and ciliary EVs is not known.

The worm is a powerful model for understanding the fundamental biology of the polycystins. The C. elegans polycystins LOV-1 and PKD-2 localize to cilia and ciliary EVs. In ciliated sensory neurons, LOV-1 and PKD-2 function in a cell-autonomous capacity to regulate male mating behaviors. In EVs, the polycystins may function non-autonomously to signal between other animals. We hypothesize that polycystin-carrying EVs may act as discrete signaling units and may carry cargoes to execute signaling, cell-targeting, and cell-uptake.

To test this idea, we used CRISPR/Cas9-mediated genome editing to generate endogenous reporters of polycystin-1 LOV-1 in C. elegans. Super-resolution Airyscan confocal microscopy was used to analyze subcellular localization and interactions of the newly generated N- and C-terminal tagged LOV-1 reporters with polycystin-2 PKD-2 in living animals. We then conducted functional assays to correlate ciliary presence of LOV-1 with ciliary function.

Our data supports that LOV-1 undergoes proteolytic cleavage at a G protein-coupled receptor proteolytic site (GPS), as mammalian polycystin-1, generating C-terminal and N-terminal fragments. We generated a functional, double-tagged version of LOV-1 (mScarlet::LOV-1::mNeonGreen), which enabled visualization of the subcellular localization of each LOV-1 fragment in the same animal. We observed that each cleavage product has different subcellular localizations. We discovered that the C-terminally tagged LOV-1 fully colocalizes with PKD-2, including its presence on ciliary EVs, whereas the N-terminally tagged LOV-1 does not. We also show that PKD-2 and C-terminal LOV-1 release on EVs is regulated i.e., not simply a waste product or a product of overexpression. In contrast, we do not observe co-localization of PKD-2 with other ciliary EV cargo, consistent with polycystin-carrying EVs acting as signaling units. We also found that LOV-1, but not PKD-2, is involved in a pathway that governs pheromone attraction of male C. elegans. Genetic analysis of this pathway suggests that ciliary presence of the N-terminal LOV-1 inhibits attraction to the pheromone. This data corroborates the notion that cleavage products of polycystin-1 might function independently, and that N-terminal LOV-1 acts as a chemosensor. Our findings will lead to better understanding of the in vivo functions of the polycystins in cilia and ciliary EVs, which may inform understanding of the human polycystins in normal and ADPKD states.

Oral presentation summaries

Molly Abbott

Institution
University of Oxford

The Surgical Management of Complications in Autosomal Dominant Polycystic Kidney Disease: A Case Study

Here I present a case of a patient with polycystic kidney disease, who unfortunately suffered several repeated hospital admissions with very severe septic infections due to the complications that arose from his kidney disease. The sheer size of them gave him a very poor quality of life, and led to these infections. Hence there was discussion by the doctors on how to best manage this, since clearly his repeated admissions meant the current management strategy of inserting stents was insufficient. This led me to research which treatment or surgical options had the best evidence supporting them. Furthermore, I discuss the difficulties he faced getting a diagnosis and since when attempting to seek care. I consider the research to suggest whether an earlier diagnosis would have reduced his complications, why he was somewhat let down by the healthcare system and the barriers he faced that led to such a late diagnosis.
Sarah Bland

Institution
University of Oklahoma Health Sciences Center

Co-authors
Alex Yashchenko and
Kurt Zimmerman

Single cell RNA sequencing uncovers a population of Trem2+ kidney resident macrophages that are associated with slowly progressing cystic kidney disease

Cystic kidney diseases affect over 12 million people worldwide and represent the third leading cause of end stage renal disease. Immune cells called macrophages have been suggested to promote disease progression in animal models. However, much of the data supporting this hypothesis comes from animal models equivalent to studying disease between ages 0-25 in humans. This does not accurately reflect age of cyst onset, or rate of cyst progression in humans, where cysts begin to form the 2nd decade of life and gradually progress until end stage renal disease occurs between the 5th-9th decade. We profiled macrophages in a slowly progressing animal model (more equivalent to human disease), a rapidly progressing disease model, and non-diseased control animals, and identified a unique population of macrophages that were only present in the slowly progressing model. We propose that these macrophages may help restrict cyst growth, and studies testing this are ongoing.
Charles DeRossi

Institution
Icahn School of Medicine at Mount Sinai

Glycosylation as a regulator of liver disease in ARPKD

Autosomal recessive polycystic kidney disease (ARPKD) is a genetic disorder characterized by the growth of fluid-filled cysts in the kidney. The liver is also affected by the same genetic mutation in ARPKD, and as such, the majority of individuals with ARPKD will also have a liver disease called congenital hepatic fibrosis (CHF). CHF is where excessive scar tissue is deposited in the liver and can disrupt normal liver function. Currently, there are no treatments to cure the liver disease in CHF/ARPKD except for liver transplantation. Therefore, more research efforts to better understand how we can prevent or reverse liver scarring in ARPKD is essential.
This project focuses on the role of glycosylation in CHF/ARPKD. Glycosylation is the process where sugars are added to target molecules, such as proteins, to direct their function or location within or outside a cell. When glycosylation is disrupted, this can lead to a myriad of diseases, including kidney and liver diseases. Mannose is a sugar similar to glucose, and the processing of mannose is a key pathway in maintaining correct glycosylation of proteins. Our research data and clinical observations stimulated us to explore the role of glycosylation and mannose metabolism as important regulators of CHF in ARPKD. With the following aims, we will test our hypotheses that abnormal glycosylation drives liver scarring in ARPKD, and that mannose supplementation can be effective at treating ARPKD-associated liver disease.
This project is relevant to PKD as it examines an extra-renal manifestation of ARPKD from a novel viewpoint – changes in liver glycosylation associated with ARPKD and examines mannose as a potential therapy for ARPKD-associated liver disease. The Impact/Significance of this project, in addition to mannose being used therapeutically, is potentially identifying novel glycoproteins that impact severity of liver disease in ARPKD which could become therapeutic targets or diagnostic markers for severity of liver disease. This proposal provides conceptual, technical and translational innovation as it focuses on the novel idea that changes in mannose metabolism and protein glycosylation drive CHF in ARPKD patients, it uses cutting-edge glycoproteomic analyses to identify key changes in the pathobiology of CHF, and it integrates findings from divergent fields which, together, introduce the novel prospect that manipulating mannose levels in the liver can improve fibrosis in ARPKD.

Page last updated June 2022

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