Neddylation inhibition reduces liver steatosis in MAFLD mice models by promoting hepatic fatty acid oxidation via DEPTOR-mTOR axis
Marina Serrano-Maciá1, Jorge Simón1.2, Maria J. González-Rellan3,4, Mikel Azkargorta5, Naroa Goikoetxea-Usandizaga1, Fernando Lopitz-Otsoa6, Diego Saenz De Urturi7, Rubén Rodríguez-Agudo1, Sofia Lachiondo-Ortega1, Maria Mercado-Gomez1, Virginia Gutiérrez de Juan6, Maider Bizkarguenaga6, David Fernández-Ramos2,6, Xabier Buque7,8, Guido A. Baselli 9,10, Luca V.C. Valenti9,10, Paula Iruzubieta11,12, Javier Crespo11,12, Erica Villa13, Jesus M. Banales7,14, Matias A. Avila7,15, Jose J.G. Marin7,16, Patricia Aspichueta6,8, James Sutherland17, Rosa Barrio17, Ugo Mayor18,19, Félix Elortza4, Dimitris P. Xirodimas20, Rubén Nogueiras 2,3, Teresa C. Delgado1*, María Luz Martínez-Chantar1,2*
1 Liver Disease Lab, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), 48160 Derio, Bizkaia, Spain. 2 Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Carlos III National Health Institute, Madrid, Spain. 3 Department of Physiology, Center for Research in Molecular Medicine and Chronic Diseases (CIMUS), University of Santiago de Compostela-Instituto de Investigación Sanitaria, Galician Agency of Innovation (GAIN), Xunta de Galicia, 15782 Santiago de Compostela, Spain. 4 Centro de Investigación Biomédica en Red de Fisiopatologia de la Obesidad y Nutrición (CIBERobn), Carlos III National Health Institute, Madrid, Spain. 5 Proteomics Platform, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Carlos III Networked Proteomics Platform (ProteoRed-ISCIII), 48160 Derio, Bizkaia, Spain. 6 Precision Medicine and Metabolism Lab, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), 48160 Derio, Bizkaia, Spain. 7 Department of Physiology, Faculty of Medicine and Nursing, University of the Basque Country UPV/EHU, 48940 Leioa, Bizkaia, Spain.8 Biocruces Bizkaia Health Research Institute, 48903 Barakaldo, Bizkaia, Spain. 9 Department of Pathophysiology and Transplantation, Università degli Studi di Milano, 20122 Milan, Italy. 10 Department of Transfusion Medicine and Hematology – Translational Medicine, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, 20122 Milan, Italy. 11 Gastroenterology and Hepatology Department, Marqués de Valdecilla University Hospital, 39008 Santander, Spain. 12 Clinical and Translational Digestive Research Group, Research Institute Marqués de Valdecilla (IDIVAL), 39011 Santander, Spain. 13 Department of Gastroenterology, Azienda Ospedaliero-Universitaria & University of Modena and Reggion Emilia, 41121 Modena, Italy. 14 Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute, Donostia University Hospital, University of the Basque Country (UPV/EHU), Ikerbasque, 20014 San Sebastian, Spain. 15 Hepatology Programme, Center for Applied Medical Research (CIMA), IDISNA, University of Navarra, 31008 Pamplona, Spain. 16 Experimental Hepatology and Drug Targeting (HEVEFARM), IBSAL, University of Salamanca, 37007 Salamanca, Spain. 17 Ubiquitin-likes And Development Lab, CIC bioGUNE, Basque Research and Technology Alliance (BRTA), 48160 Derio, Bizkaia, Spain 18 Department of Biochemistry and Molecular Biology, Faculty of Science and Technology, University of the Basque Country (UPV/EHU), 48940 Leioa, Spain. 19 Ikerbasque - Basque Foundation for Science, Bilbao, Spain).20 Centre de Recherche en Biologie cellulaire de Montpellier (CRBM-CNRS), Univ. Montpellier, UMR5237, Montpellier 34090, Cedex 5, France. Scan to download the poster 1 INTRODUCTION 4 RESULTS 4.3 mTOR inhibition via DEPTOR accumulation plays a role in neddylation inhibition 4.4 Neddylation inhibition boosts fatty acid oxidation coupled with oxidative 4.1 Hepatic neddylation is augmented in clinical and pre-clinical MAFLD Neddylation is a druggable and reversible ubiquitin-like post- mediated anti-steatotic effects in NAFLD pre-clinical models. phosphorylation in NAFLD pre-clinical models. Hepatic global neddylation levels were increased in liver biopsies from a cohort of well-characterized MAFLD patients, both lean translational modification upregulated in many diseases including To better understand MAFLD at the proteome level we performed high-throughput proteomics Liquid mTOR signaling plays an important role on regulating lipid metabolism, such as lipogenesis and and obese, in comparison to age-matched healthy controls analyzed by immunohistochemistry (IHC) (Fig. A). In-depth analysis Chromatography-Mass Spectrometry (LC-MS)-based analyses in animals models of the disease. Ingenuity oxidative fluxes in the liver. MLN4924 treatment to isolated mouse hepatocytes stimulated with OA metabolic-associated fatty liver disease (MAFLD), liver fibrosis and revealed that hepatic global neddylation levels correlate positively with the NAS score (Fig. B). Then, we evaluated the levels of pathway analysis (IPA) identified the major canonical pathways involved in NAFLD suggesting that eukaryotic induces FAO activity (Fig. O). In agreement, neddylation inhibition by MLN4924 significantly induced hepatocellular carcinoma (HCC). hepatic neddylation in mouse models of diet-induced MAFLD reflecting different stages of the pathology. In all these animal initiation factor 2 (EiF2) signaling and mTOR pathway, which plays an important role in the regulation of lipid oxidative phosphorylation (OXPHOS), an electron transport-linked phosphorylation, as well as ATP- models of MAFLD, hepatic global neddylation was induced (Fig. C-E), with higher increases corresponding to more aggressive MAFLD is a complex liver disease and comprehends a group of metabolism, are highly altered in MAFLD (Fig. K). linked respiration in mouse hepatocytes, measured by Seahorse-based analysis (Fig. P). Likewise, FAO dietary interventions. conditions being the massive accumulation of fat in the liver the activity was also shown to be induced in MLN4924-treated CDHFD and 0.1% MCDD-fed rodents (Fig. Q). A C D E K Canonical pathways Canonical pathways in e Healthy NAFLD Control HFD (20wk) Control CDHFD (6wk) Control 0.1% MCDD (4wk) O P c on Q
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+ + e Control NAFLD N N H H L .1 % L D D A A S 0 .1 C C O O S NAS Score M 0 M 3 METHOD CONCLUSIONS ACKNOWLEDGEMENTS Standard Chow Vehicle 5 6 Neddylation inhibition was evaluated in mouse isolated hepatocytes. Moreover, male adult C57BL/6 mice (3- diet Herein, we have further addressed the relevance of hepatic neddylation in MAFLD as well as the therapeutic 0 21 28 35 42 Days CONTACT INFORMATION month old) fed either with 0.1% methionine and choline deficient diet (0.1%MCD diet) or with a choline-deficient Choline-deficient high fat diet (CDHFD) (6 efficacy of neddylation inhibition both in vitro cell models and in mouse models of diet induced MAFLD. We provide weeks) Vehicle or MLN4924 (60mg/Kg bw) Standard Chow Choline-deficient high fat diet Teresa C. Delgado high fat diet (CD-HFD) were used. After 2 weeks of 0.1%MCD diet or 3 weeks of CD-HFD, mice were treated diet (CDHFD) (6 weeks) evidence that hepatic neddylation inhibition decreases liver steatosis by boosting fatty acid oxidation in a process [email protected]
Metabolism, alcohol and toxicity Marina Serrano-Macia 0 21 28 35 42 Days -21 0 21 42 Standard Chow diet during 2 or 3 more weeks, depending on the diet, with Pevonedistat (60mg/Kg) by oral gavage each 4 days. The Vehicle Days partly mediated by impaired the mammalian target of rapamycin (mTOR) signaling as regulated by DEPTOR (DEP- AAV-DIO-Control or María Luz Martínez-Chantar AAV-DIO-shNEDD8 effects of neddylation specific inhibition were also evaluated in male adult C57BL/6 AlfpCre mice infected with 0 15 18 21 24 28 Days domain containing mTOR-interacting protein). Moreover, we have identified for the first time that the levels of [email protected] 0.1 % Methionine and choline-deficient diet (0.1%MCDD) (4 weeks) NEDD8 in serum appear to correlate with NAFLD disease progression. Overall, treating NAFLD by targeting AAV-DIO-shNEED8 and maintained on CD-HFD for 6 weeks. Finally, the impact of hepatic neddylation in patients Vehicle or MLN4924 (60mg/Kg bw) Marina Serrano-Maciá neddylation may be a fast and effective strategy to regulate altered signaling pathways and metabolic reactions. [email protected] with MAFLD as well as the potential use of NEDD8 serum levels for MAFLD diagnostic purposes were evaluated. 0 15 18 21 24 28 Days
PO-2192
ILC2021