Development of an in vitro cholestatic drug-induced liver injury evaluation system using HepG2-hNTCP-C4 cells in sandwich configuration. (December 2019)

• Record Type: Journal Article
• Authors: Sakai, Yoko; Okumura, Hiroki; Iwao, Takahiro; Watashi, Koichi; Ito, Kousei; Matsunaga, Tamihide
• Is Part Of: Toxicology in vitro. Volume 61(2019)
• Publication Date: 2019-12
• Subjects: Bile acids -- Sandwich culture -- HepG2-hNTCP-C4 cells -- Drug-induced liver injury -- Na+-taurocholate cotransporting polypeptide; BAs bile acids -- BCRP breast cancer resistance protein -- BSEP bile salt export pump -- CDF 5-(and-6)-carboxy-2′, 7′-dichlorofluorescein -- CDFDA 5-(and-6)-carboxy-2′, 7′-dichlorofluorescein diacetate -- CsA cyclosporin A -- DAPI 4, 6-diamidino-2-phenylindole -- DILI drug-induced liver injury -- DMSO dimethyl sulfoxide -- FBS fetal bovine serum -- FXR farnesoid X-receptor -- GFR-Matrigel Matrigel matrix Growth Factor Reduced -- HBSS Hank's balanced salt solution -- HPHs human primary hepatocytes -- MRP multidrug resistance-associated protein -- NTCP Na+-taurocholate cotransporting polypeptide -- PBS phosphate-buffered saline -- SCHepG2-hNTCP cells sandwich-cultured HepG2-hNTCP-C4 cells -- SCHHs sandwich- cultured human hepatocytes -- tauro-nor-THCA-24-DBD N-(24-[7-(4-N, N-dimethylaminosulfonyl-2, 1, 3-benzoxadiazole)]amino-3α, 7α, 12α-trihydroxy-27-nor-5β-cholestan-26-oyl)-2′-aminoethanesulfonate; Toxicity testing -- In vitro -- Periodicals; Toxicology -- Periodicals; 615.9
• DOI: 10.1016/j.tiv.2019.104619
• Languages: English
• ISSNs: 0887-2333

Abstract

Abstract: Toxicological approaches in screening drugs that cause drug-induced liver injury (DILI) are urgently needed to reduce the risk of developing DILI and avoid immense costs resulting from late-stage drug withdrawal from clinical trials. Cholestatic DILI is characterized by bile acid (BA) accumulation in hepatocytes, typically caused by drug-induced inhibition of important bile transporters, such as bile salt export pump (BSEP) and multidrug resistance-associated protein 2/3/4 (MRP2/3/4). Therefore, NTCP expression is essential for construction of an in vitro hepatocellular toxicity evaluation system. Here, we investigated whether sandwich-cultured HepG2-hNTCP-C4 (SCHepG2-hNTCP-C4) cells were applicable for evaluation of cholestatic DILI. In SCHepG2-hNTCP-C4 cells, NTCP and MRP2/4 expression levels were comparable to those in human primary hepatocytes; however, BSEP expression was low. In addition, the substrates tauro-nor-THCA-24 DBD and CDF confirmed the functionality of NTCP and MRP2, respectively. When 22 known hepatotoxins were exposed to BAs to evaluate cholestatic DILI, cytotoxicity in SCHepG2-hNTCP-C4 cells was more frequent than that in SCHepG2 cells. Thus, SCHepG2-hNTCP-C4 cells may be useful preclinical screening tools to predict the risk of cholestatic DILI induced by drug candidates. However, further studies are needed to determine why the cholestatic cytotoxicity of some compounds would be still insufficient in SCHepG2-hNTCP-C4 cells. Graphical abstract: Unlabelled Image Highlights: Gene expression in HepG2-hNTCP-C4 cells was superior to that in HepG2 cells. HepG2-hNTCP-C4 cells could incorporate BAs into hepatocytes. We established a cholestatic DILI prediction system using SCHepG2-hNTCP-C4 cells. All hepatopathic drugs caused cholesteric cytotoxicity in SCHepG2-hNTCP-C4 cells.