Roscovitine analogues as targeted pharmacotherapies for rare cholestatic liver diseases – RAPRACLID

Bile secretion is an essential function of the liver necessary for fat digestion as well as elimination of xenobiotics and endogenous metabolites. This function mainly depends on ATP-Binding Cassette (ABC) transporters located at the canalicular membrane of hepatocytes and responsible for the secretion of hydrophobic components into bile: ABCB4 for phospholipids and ABCB11 for bile acids. Genetic variations of these transporters are associated with rare cholestatic liver diseases, the most severe being Progressive Familial Intrahepatic Cholestasis (PFIC) type 2 (defects in ABCB11) and 3 (defects in ABCB4) in which children are affected during the first months of life. While ursodeoxycholic acid has exhibited satisfying relief in most patients with mild forms of ABCB4- and ABCB11-related diseases, most of PFIC2/3 patients do not or only poorly respond to this treatment, for whom liver transplantation remains the only alternative. In order to avoid or at least delay transplantation, our research project aims at identifying new small molecules of therapeutic interest able to correct the molecular defects of ABCB4 and ABCB11 due to mutations of the genes encoding these two transporters.

Based on previous collaborative projects involving the three partners of this project, we will use structural kinase-inactive analogues of roscovitine, a trisubstituted purine, to rescue the intracellular traffic and the function of defective variants of ABCB4 and ABC11 identified in patients with rare cholestatic diseases. The ideal drug candidates must rescue the intracellular traffic without impairing transporter function. The candidate molecules will be selected using the following research approach:

1) Kinase-inactive roscovitine analogues will be synthesized by varying the chemical scaffold and substituents of the parent molecule, giving rise to a potential chemical library of more than 70,000 compounds.

2) The capacity of the newly synthesized molecules to rescue the intracellular traffic and the function of defective ABCB4 and ABCB11 genetic variants will be investigated in relevant cell models. The molecules displaying the best rescuing capacities and the less cytotoxic effects will undergo further validation in mouse models.

3) The best molecules will be chemically optimized in order to improve their benefit/toxicity ratio, by e.g., increasing efficacy at low doses. Such optimization will benefit from in silico experiments providing a realistic atomic-scaled pictures of structural defects of ABCB4 and ABCB11 variants. Particular attention will be paid to the lowering of inhibitory interactions between drug candidates and canalicular ABC transporters by means of molecular dynamics simulations.

4) Finally, the selected drug candidates will undergo a series of preclinical tests in mouse models in order to: i) determine their ADMET profile (Absorption, Distribution, Metabolism, Elimination and Toxicity); ii) validate their efficacy to rescue bile secretion in mouse models mimicking the human diseases (AAV8-mediated expression of the mutated human transporters in a KO background).

This project gathers the expertise of three partners (two academics and a private biotech company) in the fields of medicine, pharmacology, biology, biochemistry, medicinal chemistry and molecular modelling. Our main goal is to provide a solid proof-of-concept that selected roscovitine analogues could be considered as relevant pharmacological alternative to liver transplantation for patients with rare cholestatic diseases related with molecular defects of ABCB4/ABCB11. This preclinical proof-of-concept is a prerequisite for the industrial development and the marketing of new drug candidates. Moreover, new targeted pharmacotherapies identified in the frame of this project might also benefit to more diseases caused by other impaired intracellular traffic processes related with genetic variations in patients.