Generation of a bile salt export pump deficiency model using patient-specific induced pluripotent stem cell-derived hepatocyte-like cells

Abstract

Bile salt export pump (BSEP) plays an important role in hepatic secretion of bile acids and its deficiency results in severe cholestasis and liver failure. Mutation of the ABCB11 gene encoding BSEP induces BSEP deficiency and progressive familial intrahepatic cholestasis type 2 (PFIC2). Because liver transplantation remains standard treatment for PFIC2, the development of a novel therapeutic option is desired. However, a well reproducible model, which is essential for the new drug development for PFIC2, has not been established. Therefore, we attempted to establish a PFIC2 model by using iPSC technology. Human iPSCs were generated from patients with BSEP-deficiency (BD-iPSC), and were differentiated into hepatocyte-like cells (HLCs). In the BD-iPSC derived HLCs (BD-HLCs), BSEP was not expressed on the cell surface and the biliary excretion capacity was significantly impaired. We also identified a novel mutation in the 5′-untranslated region of the ABCB11 gene that led to aberrant RNA splicing in BD-HLCs. Furthermore, to evaluate the drug efficacy, BD-HLCs were treated with 4-phenylbutyrate (4PBA). The membrane BSEP expression level and the biliary excretion capacity in BD-HLCs were rescued by 4PBA treatment. In summary, we succeeded in establishing a PFIC2 model, which may be useful for its pathophysiological analysis and drug development.

Pluripotent stem cell-derived bile canaliculi-forming hepatocytes to study genetic liver diseases involving hepatocyte polarity

“…Pluripotent HUES9 cells were differentiated toward hepatocyte-like cells (hiHeps) using a protocol based on embryonic hepatic development and previous studies on hepatocyte differentiation in culture [16,24,30] ( Figure 1A). Figure S1A).…”

“…Our results suggest that curcumin and 4-phenylbutyrate may not be suitable to restore ATP7B-H1069Q trafficking in all patients and demonstrate the importance of investigating the usefulness of therapeutic compounds in the context of patientspecific, autologous ATP7B mutant proteins. 16 Copper overload has been reported in patients with MEDNIK syndrome who have no mutations in ATP7B but mutations in the AP1S1 gene [37]. Whether copper overload in MEDNIK patients is due to a ATP7B localization or trafficking defect, as suggested [34,38], has not been determined.…”

“…Patientown cells are the ideal source to study endogenous mutant proteins. The ability to generate induced pluripotent stem cells (iPSC) from patients' cells and to differentiate these to hepatocytes has been a major step towards the study of endogenous mutant proteins, including ATP7B, in physiologically relevant cell types [16][17][18][19][20][21][22][23][24]. However, the potential of iPSC-derived hepatocytes to polarize and form bile canaliculi, and their potential to study the polarized trafficking of endogenously expressed mutant protein, is not known.…”

Liver safety evaluation of endothelin receptor antagonists using HepatoPac®: A single model impact assessment on hepatocellular health, function and bile acid disposition

“…Although this approach may address issues of consistent resourcing of human reagents, maintenance of significant xenobiotic biotransformation capacity (Kratochwil et al, ) and transporter functionality related to bile salt uptake and efflux (Ulvestad et al, ) may still need to be addressed. While the notion to explore DILI causality assessments may be possible by using induced pluripotent stem cell‐derived hepatocytes from affected patients for testing, only recently has it been possible to observe loss of function genetic Single Nucleotide Polymorphisms (SNPs) for transporter functionality like BSEP expressed in culture (Imagawa et al, ). Therefore, it may be possible in the near future to truly test the impact of genetic mutations on the manifestation of drug toxicity in culture that is not the product of degradation of functionality due to harvesting/cryopreservation and culture conditions.…”

Clinical Hepatocyte Transplantation: What Is Next?

“…Furthermore, advances in gene editing technologies have facilitated the possibility of correcting the specific genetic anomaly that induce disease and subsequently engineer disease-free autologous cells for re-introduction via HT (118). These technologies are already being tested in murine xenograft models whereby human HLC are being differentiated from iPSCs, engrafted into livers of immunosuppressed mice, and then used to study physiology as well as develop new pharmacologic therapies (119, 120). …”