Caballero F.J. (Presentador) (1), Rivilla I. (2), Herraez E. (3), Briz O. (3), Santos-Laso A. (1), Izquierdo-Sánchez L. (1), Lee-Law P.Y. (1), Rodrigues P.M. (1), Jin S. (4), Gradilone S. (4), Perugorria M.J. (1), Esteller M. (5), Bujanda L. (1), Marín J.J.G. (3), Cossío F.P. (2), Banales J.M. (1) (1) Biodonostia Health Research Institute – Donostia University Hospital –, UPV/EHU, Donostia-San Sebastian, Spain. Servicio de Department of Liver and Gastrointestinal Diseases (2) Center of Innovation in Advanced Chemistry (ORFEO-CINQA), University of the Basque Country/Euskal Herriko Unibertsitatea (UPV/EHU), Donostia International Physics Center (DIPC), Donostia-San Sebastian, Spain. Servicio de Department of Organic Chemistry I (3) Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain. Servicio de Experimental Hepatology and Drug Targeting (HEVEFARM) (4) The Hormel Institute, University of Minnesota, Austin; Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota. Servicio de Department of Gastroenterology and Hepatology (5) School of Medicine and Health Sciences, University of Barcelona (UB), Barcelona, Catalonia, Spain. Servicio de Physiological Sciences Department
Polycystic liver diseases (PLDs) include a heterogeneous group of genetic disorders characterized by progressive development of multiple fluid-filled biliary cysts. We have previously shown that ursodeoxycholic acid (UDCA) inhibits hepatic cystogenesis in experimental models and patients with PLD. Furthermore, histone deacetylase 6 (HDAC6) inhibitors reduced hepatorenal cystogenesis in PCK rats. However, both therapies showed limited efficacy. Since UDCA does not display any HDAC6 inhibitory activity, and most of the available HDAC6 inhibitors are potentially toxic, we aimed to design new synthetic UDCA conjugates with selective HDAC6 inhibitory capacity (HDAC6i-UDCA) and evaluate their dual therapeutic potential for PLD.
Ten synthetic HDAC6i-UDCA#1-10 conjugates were designed, analyzed in silico, synthesized and tested for their inhibitory activity on HDAC6 and nuclear HDACs. The best five candidates were subjected to enzymatic IC50 determination on HDAC6 and HDAC1. PCK rats were orally administered 15 mg/kg day of HDAC6i-UDCA#1 for 5 months. After sacrifice, serum, bile, liver and kidney were collected and analyzed. The molecular mechanisms of HDAC6i-UDCA#1 were investigated in PLD cholangiocytes.
4 of the 10 designed HDAC6i-UDCA conjugates presented highly selective HDAC6 inhibitory activity, with HDAC6i-UDCA#1 emerging as the most promising molecule. In silico experiments predicted an active contribution of UDCA to the HDAC6 inhibitory activity of HDAC6i-UDCA#1, which was confirmed by IC50 determination. HDAC6i-UDCA#1 treatment significantly reduced liver and kidney weights, respective tissue to body weight ratios and liver cystogenesis. In parallel, increased serum levels of albumin and reduced serum levels of urea were found in treated PCK rats. Administration of HDAC6i-UDCA#1 increased the liver and kidney alpha-tubulin acetylation, a HDAC6 target, and restored the related primary cilium length in cystic cholangiocytes. This treatment also resulted in increased UDCA concentration in liver, bile, peripheral and portal blood. In vitro, HDAC6i-UDCA#1 inhibited the 2- and 3-dimentional growth of cystic cholangiocytes and decreased ERK1/2 phosphorylation, in a greater extent than its forming components, alone or in combination. HDAC6i-UDCA#1 was transported into cells through NTCP, OCT1 and OCT3 transporters.
HDAC6i-UDCA#1 inhibited hepatorenal cystogenesis in an animal model of PLD and displayed greater antiproliferative effects on PLD cholangiocytes than UDCA alone. In addition to the HDCA6 inhibitory capacity, increased UDCA concentrations in treated animals, suggest a release of UDCA upon metabolism of HDAC6i-UDCA#1, minimizing potential toxicity of metabolites and highlighting the relevance of this promising dual therapy for the treatment of PLDs.