Molecular and Clinical Links between Drug-Induced Cholestasis and Familial Intrahepatic Cholestasis

Abstract

Idiosyncratic Drug-Induced Liver Injury (iDILI) represents an actual health challenge, accounting for more than 40% of hepatitis cases in adults over 50 years and more than 50% of acute fulminant hepatic failure cases. In addition, approximately 30% of iDILI are cholestatic (drug-induced cholestasis (DIC)). The liver's metabolism and clearance of lipophilic drugs depend on their emission into the bile. Therefore, many medications cause cholestasis through their interaction with hepatic transporters. The main canalicular efflux transport proteins include: 1. the bile salt export pump (BSEP) protein (ABCB11); 2. the multidrug resistance protein-2 (MRP2, ABCC2) regulating the bile salts' independent flow by excretion of glutathione; 3. the multidrug resistance-1 protein (MDR1, ABCB1) that transports organic cations; 4. the multidrug resistance-3 protein (MDR3, ABCB4). Two of the most known proteins involved in bile acids' (BAs) metabolism and transport are BSEP and MDR3. BSEP inhibition by drugs leads to reduced BAs' secretion and their retention within hepatocytes, exiting in cholestasis, while mutations in the ABCB4 gene expose the biliary epithelium to the injurious detergent actions of BAs, thus increasing susceptibility to DIC. Herein, we review the leading molecular pathways behind the DIC, the links with the other clinical forms of familial intrahepatic cholestasis, and, finally, the main cholestasis-inducing drugs.

Keywords: ABCB1; ABCB11; ABCB4; ABCC2; BSEP protein; MDR1 protein; MDR3 protein; MRP2; drug-induced cholestasis; familial intrahepatic cholestasis; idiosyncratic drug-induced liver injury.

Figure 1

If R-value is greater than 5, a hepatocellular iDILI occurs; if R-value is less than 2, a cholestatic iDILI is diagnosed. Finally, if R-value is between 2 and 5, a mixed iDILI, cholestatic and hepatocellular, is present [5,6]. Abbreviations: ALT: alanine aminotransferase; ALP: alkaline phosphatase; ULN: upper limit of normal values; R: ratio; RUCAM: Roussel Uclaf Causality Assessment Method scale.

Figure 2

Simplified representation of the proteins expressed by the genes involved in the main pathway of synthesis, transport, and reuptake of BAs in the biliary canaliculus and hepatocyte membrane. Created with BioRender.com, accessed on 10 February 2023. The ATP-binding cassette superfamily proteins are localized in the biliary canaliculus, and mutations in these genes are associated with cholestatic liver diseases: the ABCB1 gene encodes the transmembrane transporter P-glycoprotein MDR1 that works as a drug-transport pumping out a varied range of xenobiotics from cells, and it is required for PC secretion into bile. ABCC2 synthesizes MRP2, an integral membrane glycoprotein expressed mainly in the canalicular membrane of liver cells. It transports endogenous and exogenous anionic conjugates from hepatocytes to bile. MRP2 is also involved in the resistance of cancer cells to chemotherapeutic drugs. The ABCB11 gene is responsible for BSEP synthesis; this protein transports taurocholate and other cholate conjugates from hepatocytes to the bile, resulting in the primary determinant of bile formation and bile flow. Finally, the ABCB4 gene-related protein, MDR3, is a lipid translocase or flippase causing the PC translocation to the inner leaflet of the membrane into the bile; in the absence of phospholipids such as PC, BAs cannot form mixed micelles, and the bile results are extremely hydrophobic, causing cell inflammation, bile ductular reaction and fibrosis. Abbreviations: ABCG5, ATP-binding cassette sub-family G member 5; BSEP, bile salt export pump; cAMP/GMP, cyclic adenosine monophosphate/cyclic guanosine monophosphate; FIC1, familial intrahepatic cholestasis type 1; FXR, farnesoid X receptor; MDR, multidrug resistance protein; MRP, multidrug resistance-associated protein; NTCP, sodium/taurocholate cotransporting polypeptide; OATP, organic anion-transporting polypeptide; TJP2, tight junction protein 2.

Figure 3

Molecular visualization of likely benign variant p.S893A in ABCB1 (MDR1 protein) by using PyMOL, accessed on 30 January 2023. The residue change of amino acid serine to alanine causes two bond losses for –OH groups with the other two near residues in the helix. Transcript reference: ENST00000622132.5.

Figure 4

AlphaFold structure prediction of ATP-binding cassette sub-family C member 2. All five reported variations for ABCC2 were mapped on available 3D structures, as assessed through UniProt. AF-Q92887-F1-model_v4 of ABCC2 protein in helix, turn, and beta strand conformation. Visualization of the protein and residues has been performed by PyMOL (https://pymol.org/2/, accessed on 30 January 2023). Here, residues are shown in the text: p.H1496H (c.4488C>T) and p.C1515Y (c.4544G>A) (ABCC2 NM_000392.5). Residues are labeled as colored spheres according to their actual classification on Varsome. P/LP, red spheres: pathogenic/likely pathogenic; VUS, orange spheres: variant at uncertain significance; B/LB, blue spheres: benign/likely benign.

Figure 5

Human bile salt exporter ABCB11 in complex with taurocholate. All 12 reported variations for ABCB11 were mapped on available 3D structures, as assessed through UniProt. Protein Data Bank (PDB) code 7e1a of BSEP protein in helix, turn, and beta strand conformation. Visualization of the protein and residues has been performed by PyMOL (https://pymol.org/2/, accessed on 30 January 2023). Residues are labeled as colored spheres according to their actual classification on Varsome [24].

Figure 6

Structure of phosphatidylcholine translocator ABCB4. Cryo-EM structure of nanodisc-reconstituted human ABCB4 trapped in an ATP-bound state at a resolution of 3.2 Å. The nucleotide-binding domains form a closed conformation containing two bound ATP molecules, but only one of the ATPase sites has bound Mg²⁺ (dark grey). The transmembrane domains adopt a collapsed conformation at the level of the lipid bilayer. All 12 reported variations for ABCB4 were mapped on available 3D structures, as assessed through UniProt. PDB code 6s7p of MDR3 protein in helix, turn, and beta-strand conformation. The spatial position of mutation R652G (single sphere) is inferred because no 3D structure was available for residues from 629 to 692 (magenta). The protein is in a lipid environment with a cholesterol structure in this representation. Visualization of the protein and residues has been performed by PyMOL (https://pymol.org/2/, accessed on 30 January 2023). Residues are labeled as colored spheres according to their actual classification on Varsome [26].