Academic literature on the topic 'Pfic2'

ABCB11/BSEP (Bile Salt Export Pump) est exprimé à la membrane canaliculaire des hépatocytes. Sa fonction de transport d’acides biliaires dans la bile est essentielle à la sécrétion biliaire. Près de 400 variations du gène ABCB11 ont été identifiées et sont associées à des maladies hépatobiliaires rares, la plus sévère étant la cholestase intrahépatique progressive familiale de type 2 (PFIC2). L’efficacité des traitements médicaux est limitée. Par conséquent, une transplantation hépatique est indiquée avant l’âge adulte pour près de deux tiers des patients. Dans ce contexte, l’identification de thérapies alternatives est un enjeu capital.Cette thèse s’intéresse à la recherche de stratégies thérapeutiques personnalisées permettant de corriger les conséquences pathologiques de certaines variations d’ABCB11 identifiées chez des patients. Dans le cadre d’une stratégie de traitement par des molécules potentiatrices, nous avons étudié les variations A257V, G562D et T463I d’ABCB11 par modélisation moléculaire 3D. L’étude de l’expression et de la fonction de ces variants dans différents modèles cellulaires a confirmé que ces variations étaient responsables d’un défaut de fonction du transporteur Abcb11. L’ivacaftor (VX 770, Kalydeco®), approuvé cliniquement pour le traitement de la mucoviscidose, corrigeait le défaut d’activité de ces trois variants.Des effets similaires ont été observés avec les molécules GLPG1837, SBC040 et SBC219, connues comme potentiateurs de CFTR (Cystic Fibrosis Transmembrane conductance Regulator). Dans une optique de thérapie combinatoire, nous avons également mis en évidence la capacité de ces potentiateurs à corriger le défaut de fonction des variants R1090C et R1090W, produits potentiels de la translecture du variant non-sens R1090X. Nous avons également évalué les molécules correctrices elexacaftor (VX-445) et tezacaftor (VX-661). Ces correcteurs, en monothérapie ou en combinaison, permettaient de restaurer l’adressage du variant R1128C, s’accompagnant d’une augmentation significative du transport de taurocholate. De façon intéressante, l’addition de molécules potentiatrices réduisait ces effets.L’ensemble de ces travaux constitue une preuve de concept que les défauts de certains variants d’ABCB11 peuvent être corrigés par ces molécules potentiatrices à haut potentiel thérapeutique. Ce type de traitements pourrait être envisagé pour les patients atteints de déficit en ABCB11 et permettrait ainsi d’augmenter la pharmacopée disponible pour traiter ce genre de pathologies et ainsi repousser voir palier à la transplantation hépatique pour les cas les plus sévères<br>ABCB11/BSEP (Bile Salt Export Pump) is expressed at the canalicular membrane of hepatocytes. It ensures bile acids secretion into bile which is essential for biliary secretion. Nearly 400 variations of the ABCB11 gene have been identified and are associated with rare hepatobiliary diseases, the most severe being progressive familial intrahepatic cholestasis type 2 (PFIC2). The effectiveness of medical treatments is limited. Consequently, liver transplantation is required before adulthood for almost 2/3 of PFIC2 patients. In this context, the identification of alternative therapies is a major challenge.This thesis focuses on personalized therapeutic strategies to correct the pathological consequences of some ABCB11 variations identified in patients. The A257V, G562D and T463I variations of ABCB11 were studied by 3D molecular modelling. These variations were responsible for a defect in Abcb11 transport function. Ivacaftor (VX-770, Kalydeco®), a clinically approved cystic fibrosis treatment, corrects the activity defect of the three variants.Similar effects were observed with GLPG1837, SBC040 and SBC219, known as potentiators of CFTR (Cystic Fibrosis Transmembrane Conductance Regulator).From a combinatory therapy perspective, we also demonstrated the ability of these potentiators to correct the transport defect of the R1090C and R1090W variants, potential readthrough products of the R1090X nonsense variant. We also evaluated the ability of Elexacaftor (VX-445) and Tezacaftor (VX 661) correctors of CFTR. These correctors, alone or in combination, restored trafficking of the R1128C missense variant, leading to a significant increase in the transport function. Interestingly, the addition of potentiators abolishes this effect.Altogether, this thesis constitutes a proof of concept that molecules with high therapeutic potential can correct the molecular defects of ABCB11 variants. These treatments could increase the pharmacopoeia available for patients with ABCB11 deficiency and thus delay or even suppress the need for liver transplantation

ABCB4/MDR3 est le transporteur canaliculaire de la phosphatidylcholine. Il est exprimé à la membrane canaliculaire des hépatocytes et est essentiel à la sécrétion biliaire. Un défaut d’ABCB4 entraîne des pathologies hépatobiliaires, dont la PFIC3 (cholestase intrahépatique familiale progressive de type 3), caractérisée par une cholestase précoce qui progresse vers la cirrhose et l’insuffisance hépatique avant l’âge adulte. Dans la majorité des cas, la seule thérapie efficace est la transplantation hépatique. Cette thèse s’intéresse aux rôles de l’extrémité C-terminale dans la régulation de la stabilité et l’expression de ce transporteur au canalicule biliaire. Nous avons délété le motif Q-N-L de cette extrémité et montré que cette délétion affecte la stabilité d’ABCB4/MDR3 en augmentant son endocytose. Son interaction avec des protéines à domaines PDZ est alors étudiée. Nous avons montré une interaction par le motif Q-N-L avec la protéine à domaines PDZ, EBP50. Cette interaction est nécessaire pour l’expression canaliculaire et la stabilité du transporteur<br>ABCB4 is a phosphatidylcholine translocator specifically expressed at the bile canalicular membrane of hepatocytes. Mutations of the ABCB4 gene cause progressive familial intrahepatic cholestasis type 3 (PFIC3), a rare genetic disease characterized by early onset of cholestasis and evolution to cirrhosis and liver failure before adulthood. Little is known regarding the molecular mechanisms which control the canalicular expression and membrane stabilization of ABCB4 in hepatocytes. The aim of this work was to study the role of the C-terminal domain of ABCB4 for its expression and stability. potential interaction with EBP50, a PDZ protein highly expressed in hepatocytes. The experimental approach consisted in the deletion of the QNL motif at the C-terminus of ABCB4. The truncation of the QNL motif leds to a reduction of ABCB4 stability by increasing its endocytosis. ABCB4 co-precipitated with EBP50, an interaction that required the QNL motif. This interaction plays a critical role in the canalicular expression and stabilization of ABCB4

Among contemporary gene transfer vehicles, non-pathogenic recombinant adeno-associated viral vectors (rAAV) show exceptional promise for liver-targeted therapeutic approaches. The broad focus of studies described in this thesis was the development of rAAV-mediated gene therapy to treat Progressive Familial Intrahepatic Cholestasis type 3. This autosomal recessive condition, caused by mutations of ABCB4, results in deficient hepatocanalicular phosphatidylcholine translocation and leads to progressive cholestatic liver disease with approximately 50% of patients requiring liver transplantation before reaching adulthood. Using an Abcb4-knockout mouse model, in vivo liver transduction with rAAV2/8 vectors encoding hABCB4 led to increased biliary phosphatidylcholine in disease-free heterozygous, but not in homozygous adults with established liver disease, despite varying vector genome size and routes of administration. Maximal transduction was achieved prior to onset of liver disease, optimally in neonates. However, loss of transgene expression occurs following neonatal vector delivery, due to rAAV episomal degradation during rapid liver growth. A novel, hybrid rAAV-piggyBac transposon vector strategy was devised to sustain hABCB4 expression in neonatally-treated homozygotes. Successful correction of liver disease was demonstrated in Abcb4-/- mice up to 9 months post-inoculation, with preliminary results indicating reduction in disease-related hepatocarcinogenic risk. These results demonstrate that rAAV-mediated gene therapy has the potential to offer patients with this heritable cholestatic liver disease an effective alternative treatment to liver transplantation, but also illustrate the importance of addressing challenges, such as the impact of liver pathology on vector performance, which is vital before this potential can be realised for this and related conditions.

La colestasi intraepatica è una sindrome clinica e bioumorale secondaria a disturbi congeniti o acquisiti della formazione e secrezione canalicolare degli acidi biliari. Le colestasi progressive familiari intraepatiche (PFIC) rappresentano la causa di colestasi nel 10-15% dei casi e mutazioni in tre geni sono state classicamente identificate: le PFIC tipo 1 e 2, causate da mutazioni nei geni ATP8B1 e ABCB11, caratterizzate da livelli normali di GGT e la PFIC di tipo 3, causata da mutazioni nel gene ABCB4, con GGT elevate. Negli ultimi anni, le tecniche di Next Generation Sequencing hanno permesso di identificare e caratterizzare nuovi geni coinvolti nelle colestasi ereditarie sia nella forme a GGT normali (TJP2, FXR, MYO5B), che elevate (CLDN1, DCDC2). A partire da tali premesse, è stato condotto uno studio multicentrico, retrospettivo e prospettico, con l’obiettivo di arruolare pazienti affetti da colestasi e caratterizzarli fenotipicamente e genotipicamente. I pazienti sono stati studiati mediante sequenziamento dell’esoma e conferma con tecnica Sanger nei probandi e, qualora disponibili, nei genitori. Sono stati arruolati 25 pazienti con i seguenti risultati: una predominanza del sesso maschile (19/25, 76%) ed un’età mediana all’esordio di 0.8 anni (range 0 - 15.1). Il 68% dei pazienti (17/25) era di origine italiana. La durata media del follow-up è stata di 3.7 anni (range 0.1 - 25.6). Le manifestazioni cliniche all’esordio comprendevano epatomegalia (68%), ittero (57%), prurito (43%), scarsa crescita (33%), feci ipocoliche (33%) e splenomegalia (28%). Il valore mediano delle ALT era 184 U/L (range 54 - 650), della bilirubina totale e diretta rispettivamente 5.5 mg/dl e 3.2 mg/dl. Gli acidi biliari erano aumentati nell’81% dei pazienti. Una lieve predominanza delle forme a GGT elevate (64%) è stata riscontrata. La biopsia epatica, eseguita in 16/25 (64%) pazienti, mostrava un’ istologia compatibile con colestasi intraepatica nelle forme a GGT normali e con colangiopatia per le forme a GGT elevate.La terapia medica all’esordio consisteva nell’utilizzo di acido ursodesossicolico, supplementazione con vitamine liposolubili e trattamento del prurito. Quattro pazienti (16%) hanno ricevuto trapianto epatico. I risultati completi delle analisi genetiche, disponibili per 8 pazienti, hanno mostrato una prevalenza di mutazioni a carico del gene della miosina 5B (MYO5B), presenti in 6 pazienti; 2 pazienti avevano mutazioni a carico del gene ABCB4 (PFIC tipo 3), mentre per i restanti pazienti le analisi genetiche sono ancora in corso.