AbstractUGT1A gene family members encode for phase II metabolic enzymes that play a crucial role in the biotransformation of endogenous and exogenous compounds into highly active, lower activity or inactive compounds that are easily excreted from the body. Many of these compounds include clinically administered drugs, hormones, bile acids and environmental toxins. Compromised or excess expression of the UGT1A genes is commonly associated with adverse consequences such as hyperbilirubinemia, cancer progression or undesired drug-drug interactions. Whilst a substantial amount of research has linked hepatic UGT1A gene expression to activated pregnane X receptor (PXR), upcoming research has evidenced extrahepatic UGT1A activity as very clinically relevant, although influenced by vitamin D receptor (VDR). Research in this direction is still at its infancy, the extent, clinical consequences and molecular mechanisms are not well understood. This project aims to characterize VDR in the detoxification processes, particularly, the regulation of UGT1A gene family members and the molecular mechanisms involved. Therefore, to address this issue, LS180 cells, that imitate 1,25D’s colonic physiological responses were used to investigate the regulation of the entire family at mRNA, protein and functional level. A series of reporter-based assays were implemented to define a functional and VDR specific binding motif. Upon identification of UGT1A4 as the most responsive to VDR ligands, novel tools such as CRISPRi genome editing, molecular cloning and characterization of the UGT1A promoter region were employed to examine putative binding motifs. To extend upon this study, novel molecular mechanisms, to investigate the reciprocal effects of cross-talk between VDR and NRF2 signalling pathways were examined. This interplay has direct implications for a range of physiological and pathological consequences, including enhanced detoxification, cancer prevention and anti-aging properties. Contrary to previous findings, we identified that VDR was in this case dependant upon intact NRF2 signalling. Furthermore, the interaction of both NRF2 and VDR signalling pathways did not significantly enhance UGT1A gene expression, although, surprisingly, inhibitory effects were observed. Although LS180 cells were predominantly used in our novel approaches, future studies should incooperate other cell model systems were NRF2 expression is abundant.
Collectively this study contributes towards our understanding of VDR and its cooperative activities that influence UGT1A gene expression and NRF2 signalling pathways. Whilst the study highlights the impact of VDR ligand co-administration with glucuronidation susceptible drugs, this study reinforces the importance of maintaining optimal 1,25D levels for chronic disease prevention, management of hereditary hyperbilirubinemia and neonatal jaundice. Perhaps, employing novel approaches using these findings will be the best advancement, where UGT1A expression is compromised.
|Date of Award
|Paul Thompson (Supervisor) & Colum Walsh (Supervisor)