Genetically Edited Hepatic Cells Expressing the NTCP-S267F Variant are Resistant to Hepatitis B Virus Infection

Abstract The sodium-dependent taurocholate co-transporting polypeptide (NTCP)-S267F variant is known to be associated with a reduced risk of HBV infection and disease progression. The NTCP-S267F variant displays diminished function in mediating HBV entry, but its function in HBV infection has not been fully established in more biologically relevant models. We introduced NTCP-S267F variant and tested the infectivity by HBV in genetically edited hepatic cells. HepG2-NTCP clones with both homozygous and heterozygous variants were identified after CRISPR base editing. NTCP-S267F homozygous clones did not support HBV infection. The heterozygotes clones behaved similarly to wild type clones. We generated genetically edited human stem cells with the NTCP-S267F variant, which differentiated equally well as wild type into hepatocyte like cells (HLC) expressing high levels of hepatocyte differentiation markers. We confirmed that HLC with homozygous variant did not support HBV infection and heterozygous variant clones were infected with HBV equally as well as the wild-type cells. In conclusion, we successfully introduce the S267F variant by CRISPR base editor into the NTCP/SLC10A gene of hepatocytes, and showed the variant is a loss-of-function mutation. This technology of studying genetic variants and their pathogenesis in a natural context is potentially valuable for therapeutic intervention against HBV.