Investigation of urinary extracellular vesicles as novel and safe therapeutics for autosomal recessive polycystic kidney disease

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ABSTRACT Autosomal recessive polycystic kidney disease (ARPKD) is a leading cause of kidney failure in childhood and is caused primarily by mutations in the PKHD1 gene as well as mutations in CYS1. Although ARPKD patients progress to end stage kidney disease at varying ages, 20-30% of ARPKD patients have severe disease which results in death 24-48 hours after birth. Despite the tremendous unmet need, limited progress has been made for ARPKD treatment and no FDA-approved therapies are available for ARPKD patients. Although tolvaptan, a small molecule drug that was approved in 2018 for autosomal dominant PKD which mostly affects adults, is currently under phase I clinical trials for ARPKD, tolvaptan is not kidney-targeted and results in off-target effects. Additionally, tolvaptan’s long-term efficacy, tolerability, and developmental consequences in a primarily pediatric ARPKD patient population and its ability to enhance survival in neonates with severe ARPKD is unknown. Thus, new therapeutic strategies that can address both safety and efficacy in ARPKD are urgently needed. To that end, the purpose of this proposal is to examine the potential of urinary extracellular vesicles (uEVs) as a novel and safe therapy for ARPKD. EVs, also known as exosomes, are secreted, membrane-bound, biological nanoparticles that facilitate cell-to-cell communication and contain RNA and protein cargo characteristic to their parent cell. We propose uEVs for ARPKD therapy for several reasons: uEVs are 1) inherently biocompatible which is critically important when developing therapies for pediatric patients, 2) carry fibrocystin and cystin, the gene products of PKHD1 and CYS1 which are mutated in ARPKD, respectively, 3) have been found to home to the kidneys and transfer functional proteins to induce a therapeutic outcome in kidney disease, and 4) obtaining high quantities uEV from urine, which is normally discarded, is feasible, noninvasive, and cheap. We hypothesize that uEVs derived from non-disease sources are a safe therapy that can be used to deliver and supplement functional proteins including fibrocystin and cystin that are defective in ARPKD to inhibit disease progression. To test our hypothesis, we will first characterize the nanoparticle properties of uEVs and evaluate uEV fibrocystin and cystin protein and mRNA cargo, cell internalization, and therapeutic effects in renal cells in vitro (Aim 1). Next, we will administer uEVs intravenously in slowly progressing and severe ARPKD murine models and evaluate the pharmacokinetic properties, therapeutic efficacy, and safety in vivo (Aim 2.1). Finally, given most human ARPKD leads to death in utero or in newborns shortly after birth, we will deliver uEVs in utero in pregnant mice and evaluate the ability to extend survival in severe ARPKD upon fetal delivery (Aim 2.2). Through our multidisciplinary, investigative team of nanomedicine and pediatric nephrology, we are well-equipped and fully committed to successfully carry out this work that is much needed in ARPKD therapy.