Group Leader Hannover Medical School Hannover, Niedersachsen, Germany
Abstract: Progressive familial intrahepatic cholestasis (PFIC) represents a group of rare disorders characterized by defective bile formation and account for 10-15% of neonatal cholestasis. The underlying mechanism of intrahepatic cholestasis can be associated with mutations in hepatobiliary transport proteins or in nuclear receptors responsible for proper transcriptional control of these transporters. In our previous work we demonstrated that patient-specific iPS cell lines can be differentiated into hepatic organoids for functional studies of the patients’ mutations. In this study, we investigated, if such organoids could serve as model to assess effective mRNA-based gene therapy. First, we studied iPSC-based organoids from two patients suffering from BSEP deficiency caused by mutations in the farnesoid-X receptor (FXR) gene NR1H4. The NR1H4 gene encodes four different FXR variants whereas the FXR alpha-2 variant is most abundant in the liver. We used one cell line carrying a homozygous variation in exon 7, resulting in a loss of substrate binding of FXR and therefore in a complete abrogation of downstream signal transduction. A second cell line carries a homozygous variation in intron 4 resulting in an exon-skipping which encodes the start codon for both FXR alpha-1 and -2 variants, while transcription of the FXR alpha-3 and -4 variants would be unaffected. We generated lipid nanoparticles (LNP) carrying the mRNA of the four different FXR variants as well as BSEP as a positive control. After treatment of the hepatic organoids with the different LNP, we were able to restore BSEP-mediated hepatobiliary transport with all FXR variants. This indicates that even the gut-associated FXR alpha-3 and -4 variants are able to restore hepatic function and might be targetable to induce BSEP function in FXR alpha-1 or -2 deficient patients. However, as the different FXR variants are involved in several metabolic pathways, the global impact of this alternative activation needs further investigation.
Funding Source: This research was funded by the German Ministry of Education and Research (BMBF) through grants to TC, UB, and VK (HIChol: 01GM2204A, 01GM2204B, 01GM2204C).
