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Urine sample-derived human induced pluripotent stem cells as a model to study PCSK9-mediated autosomal dominant hypercholesterolemia


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Human induced pluripotent stem cells (hiPSC) are becoming a relevant model for the study of liver metabolic diseases once differentiated into hepatocyte-like cells (HLC), and it has been shown that they can faithfully recapitulate autosomal dominant hypercholesterolemia (ADH). PCSK9 is a critical modulator of cholesterol homeostasis, and quickly became a hot target for ADH pharmacological treatment strategies. However, current cellular models to further decipher the role of PCSK9 in ADH are limited, especially to study the PCSK9 gain of function mutation S127R, which seems to interfere with LDL cholesterol homeostasis intracellularly by still unknown mechanisms.

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Urine sample-derived human induced pluripotent stem cells as a model to study PCSK9-mediated autosomal dominant hypercholesterolemia

  1. 1. l’institut du thorax, unité Inserm UMR 1087-CNRS UMR 6291, IRS-UN, 8 quai Moncousu, BP 70721, 44007 Nantes Cedex 1, France Transatlantic network of Excellence 2014-2019 The proprotein convertase subtilisin/kexin type 9 PCSK9 PCSK9 Mutations : Gain-of-function -> Hypercholesterolemia Loss-of-function -> Low level of circulant cholesterol, cardioprotector effects -> Development of a new class of cholesterol-lowering drug R104C/V114A (LOF) S127R (GOF) PCSK9 protein structure Schulz & Schlüter Basic Res Cardiol (2015) 110:4 1 Episomal reprogramming of UCells isolated from a healthy donor and the patient carrying the PCSK9-S127R Gain of function mutation UCells UhiPSCs 2 UhiPS cells differentiate into hepatocytes recapitulate PCSK9-GOF and LOF features in vitro3 Karim Si-Tayeb1, Méryl Roudaut1,2, Amandine Caillaud1, Aurore Girardeau1, Matthieu Pichelin1, Lucie Arnaud1, Cédric Le May1, Nathalie Maubon2, Bertrand Cariou1 1L’institut du thorax, INSERM, CHU Nantes, Nantes France 2HCS Pharma, Lille, France 5 3D hepatic differentiation of UhiPS cells enhanced the expression of genes involved in cholesterol metabolism regulation 3D scaffolds formed by crosslinking of Hyaluronic Acid with ADH (Adipic Acid Di- Hydrazide) to form reticulated chains. Control and PCSK9-S127R hepatocytes responded to pravastatin treatments and mutated cells displayed a stronger response in accordance with related patients clinical responses Gene expression 4 Functional tests Si-Tayeb, Lemaigre & Duncan Developmental Cell 2010 Differentiation - Cytotoxicity (necrosis/apoptosis) - Genotoxicity (gH2AX, comet…) - Hepatoxicity (phospholipidosis, steatosis, cholestasis…) - Target of interest (expression, localisation and trafficking) - Phenotypic modification (stress, proliferation, differenciation, …) - Lipid (LDL-C, TG) - Glucid metabolism - ADME (CYP, UGT, transporters activities…) PharmacologyToxicology Metabolism 6 HIGH CONTENT SCREENING (HCS) – PHENOTYPIC SCREENING Ongoing hiPSC-to-hepatocyte 3D-differentiation protocole setup 2(-DeltaCt)2(-DeltaCt)2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt) 2(-DeltaCt)2(-DeltaCt) High throuput differentiation