Liver infections may not make headlines like the flu or COVID-19, but they quietly impact hundreds of millions of people worldwide. Hepatitis B and C viruses (HBV and HCV) are masters of persistence, often flying under the radar for years while gradually damaging the liver. Left untreated, these infections can lead to cirrhosis, liver cancer, and the need for transplantation.
Although antiviral treatments exist, they don’t work for everyone—and in the case of HBV, they rarely offer a true cure. That’s why researchers are turning to gene therapy for fresh solutions. However, developing gene-based therapies to treat viral liver disease requires effective delivery of therapeutic molecules to human liver cells. Since these viruses replicate only in human hepatocytes, liver-humanized mouse models—engineered to contain human hepatocytes—have become essential for studying infection and testing new treatments.
In a new study published in Molecular Therapy: Methods & Clinical Development, members of Dr. Keith Jerome’s lab tested three different lipid nanoparticle (LNP) configurations, with the goal of demonstrating the preclinical utility of their previously described chimeric NSG-PiZ mouse model. A collaborative effort between principal staff scientist Dr. Daniel Stone, postdoctoral fellow Dr. Harrison Dulin and research technician Michelle Loprieno found that while LNPs injected into mice effectively targeted mouse hepatocytes, the same particles were much less effective at targeting the human hepatocytes within the model.
This was confounding, given that the same particles had already been proven to work in a non-human primate model. It was as if the delivery trucks (LNPs) were able to reach and unload their packages easily at the mouse “houses” (hepatocytes), but when they arrived at the human “houses” within the same neighborhood, they found locked doors and no way to get inside.
“Initially, Michelle and I were left frustrated after screening different LNP formulations for their ability to transfect human hepatocytes in chimeric NSG-PiZ and FRG mice [another humanized liver model]. Our ultimate goal was to deliver therapeutic mRNAs to HBV infected human hepatocytes in these mice, but we found that 3 different LNP formulations could transfect only mouse hepatocytes when delivered intravenously,” shared Stone.
Despite transient increases in liver enzymes (ALT and AST) and mild hematopoietic changes following LNP administration, both NSG-PiZ and FRG mice tolerated the LNP treatment well. Single-cell RNA sequencing demonstrated that key receptors involved in LNP uptake—including ApoE, LDL receptor, and asialoglycoprotein receptors—are expressed at comparable levels on human and mouse hepatocytes, suggesting that receptor expression differences do not fully explain the poor uptake in human cells.
Stone and Loprieno performed the initial LNP testing, and according to Stone, the group was able to crack the problem of human hepatocyte transfection wide open once Dulin joined the team. “After joining the lab, I was interested in figuring out why LNPs were only transfecting the mouse hepatocytes but not the human hepatocytes in chimeric mice. We hypothesized that serum factors that bind to LNPs might be playing a role in the targeting of LNPs to different cell types. Our data showed that serum factors from our chimeric PiZ mice could inhibit transfection of human hepatocytes in vitro. This is what led us to choose the TriGalNac LNP formulation, which can directly target the hepatocytes rather than relying on binding to serum factors for cell targeting,” shares Dulin.
TriGalNAc (short for triantennary N-acetylgalactosamine) is a synthetic, trivalent ligand composed of three N-acetylgalactosamine (GalNAc) sugar molecules arranged around a central scaffold. It is specifically designed to target the asialoglycoprotein receptor (ASGPR) on hepatocytes. Once bound, the receptor–ligand complex is endocytosed, bringing attached molecules (such as siRNA, antisense oligos, or peptides) into the cell. Using this hepatocyte-targeting molecule has enabled the group to overcome barriers in mouse serum preventing transfection of the targeted human hepatocytes, thus improving the precision and potency of their gene delivery system.
This work is yet another example of a fruitful collaboration not just between members of the Jerome lab, but also between the Jerome lab and Massachusetts-based biotechnology company Excision Biotherapeutics, who according to Loprieno are hard at work “developing novel CRISPR-based antiviral therapeutics for HBV, HIV and HSV.”
Dr. Keith Jerome shared his thoughts about the impact of their findings. “These studies represent another important step toward our goal of a curative gene therapy for chronic hepatitis B infection. We previously developed the liver humanized NSG-PiZ mouse model as an alternative to previous, more complex models of HBV, and now we’ve leveraged this model to optimize gene therapy delivery to human hepatocytes. I am incredibly excited as we move toward using the TriGalNac LNPs for delivery of actual anti-HBV therapies. The ability to cure HBV infections could potentially prevent millions of new cases of hepatocellular carcinoma, cirrhosis, and other serious diseases,” says Jerome.
Fred Hutch/University of Washington/Seattle Children’s Cancer Consortium Member Dr. Keith Jerome contributed to this research.
The spotlighted research was funded by Excision Biotherapeutics and the National Institutes of Health.
Stone D, Takeuchi R, Dulin H, Loprieno MA, Strongin DE, Saraswathi S, Cradick TJ, Aubert M, Roychoudhury P, Gordon J, Jerome KR. 2025. Serum factors create species-specific barriers to hepatic gene transfer by lipid nanoparticles in liver-humanized mice. Molecular Therapy: Methods & Clinical Development. DOI: 10.1016/j.omtm.2025.101470
Science Spotlight writer Jenny Waters is a postdoctoral research fellow in the Hsieh lab at Fred Hutch. She studies how mRNA translation coordinates bladder cancer transformation and metastasis by post-transcriptionally regulating expression of oncogenic proteins. Outside of the lab, Jenny enjoys spending time with her dogs, convincing her husband to join her on trail runs, and pretending every steep hill is just a "gentle incline."
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