Modeling Progressive Familial Intrahepatic Cholestasis Type I Caused by ATP8B1 deficiency

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PROJECT SUMMARY/ABSTRACT ATP8B1/FIC1 belongs to the P4 subfamily of P-type adenosine triphosphatases. In the liver, it is localized on the bile canalicular membrane to maintain its lipid asymmetry that is essential for proper function of the bile salt transporters. In humans, mutations in ATP8B1 result in rare forms of chronic intrahepatic cholestasis with autosomal recessive inheritance. The most severe form of ATP8B1 deficiency, progressive familial intrahepatic cholestasis type I (PFIC1), is characterized by normal serum GGT cholestasis and pruritus in early childhood that can progress into end-stage liver disease. Due to the limitations of existing experimental models, ATP8B1 is understudied and our understanding of how ATP8B1 deficiency results in cholestasis remains incomplete, precluding the development of effective therapy for PFIC1. Our long-term goal is to understand the etiology of chronic intrahepatic cholestasis and improve patient diagnosis and treatment. The overall objective for this ap- plication is to establish a novel zebrafish model for ATP8B1 deficiency to better understand the pathogenesis of cholestasis and discover new therapeutic strategies. Our preliminary study demonstrated that Atp8b1- deficient zebrafish developed cholestasis and were lethal by 2 weeks of age, providing proof of concept that zebrafish can be used to model PFIC1. In this application, we propose to conduct further phenotypic analysis of atp8b1 germline mutant zebrafish and develop assays to study ATP8B1 function in vivo. We will determine the feasibility of using zebrafish to validate patient variants and test the effect of pharmaceutical intervention on cholestasis. The rationale is that developing additional animal models of ATP8B1 deficiency will complement the existing in vitro and rodent models and bring new knowledge into our understanding of ATP8B1 function and the mechanisms of cholestasis caused by ATP8B1 deficiency. In Aim 1, we will characterize the cholestat- ic phenotypes caused by Atp8b1 deficiency in zebrafish. We will track the subcellular distribution of fluorescent lipids in the liver and intestine of wildtype and atp8b1 mutant larvae to understand how ATP8B1 regulates the transport of lipids in vivo. In Aim 2, we will determine the feasibility of using zebrafish to validate the biological function of ATP8B1 patient variants. We will introduce two common ATP8B1 missense variants into zebrafish and determine if they change ATP8B1 expression and cause cholestasis. We will test if treatment with 4- phenylbutyric acid rescues these animals. The research proposed in this application is highly innovative be- cause it establishes a novel zebrafish model of ATP8B1 deficiency that offers unique advantages for studying bile homeostasis and cholestatic phenotypes. We will also develop a pipeline for future drug screen to identify compounds that rescue cholestasis caused by ATP8B1 deficiency. The study is also significant because this new in vivo model is expected to advance our understanding of ATP8B1 function in bile homeostasis and pathogenesis of PFIC1. The results from the study will provide critical preliminary data for a highly competitive R01 application to further pursue disease mechanisms and drug discovery.