A universal system to select gene-modified hepatocytes in vivo

Sean Nygaard, Adi Barzel, Annelise Haft, Angela Major, Milton Finegold, Mark A. Kay, Markus Grompe

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Many genetic and acquired liver disorders are amenable to gene and/or cell therapy. However, the efficiencies of cell engraftment and stable genetic modification are low and often subtherapeutic. In particular, targeted gene modifications from homologous recombination are rare events. These obstacles could be overcome if hepatocytes that have undergone genetic modification were to be selectively amplified or expanded. We describe a universally applicable system for in vivo selection and expansion of gene-modified hepatocytes in any genetic background. In this system, the therapeutic transgene is coexpressed with a short hairpin RNA (shRNA) that confers modified hepatocytes with resistance to drug-induced toxicity. An shRNA against the tyrosine catabolic enzyme 4-OHphenylpyruvate dioxygenase protected hepatocytes from 4-[(2-carboxyethyl)-hydroxyphosphinyl]-3-oxobutyrate, a small-molecule inhibitor of fumarylacetoacetate hydrolase. To select for specific gene targeting events, the protective shRNA was embedded in a microRNA and inserted into a recombinant adeno-associated viral vector designed to integrate site-specifically into the highly active albumin locus. After selection of the gene-targeted cells, transgene expression increased 10- to 1000-fold, reaching supraphysiological levels of human factor 9 protein (50,000 ng/ml) in mice. This drug resistance system can be used to achieve therapeutically relevant transgene levels in hepatocytes in any setting.

Original languageEnglish (US)
Article number342ra79
JournalScience Translational Medicine
Volume8
Issue number342
DOIs
StatePublished - Jun 8 2016

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Hepatocytes
Transgenes
Small Interfering RNA
Genes
Dioxygenases
Gene Targeting
Homologous Recombination
Cell- and Tissue-Based Therapy
Drug-Related Side Effects and Adverse Reactions
MicroRNAs
Drug Resistance
Genetic Therapy
Tyrosine
Albumins
Liver
Enzymes
Proteins
Therapeutics

ASJC Scopus subject areas

  • Medicine(all)

Cite this

Nygaard, S., Barzel, A., Haft, A., Major, A., Finegold, M., Kay, M. A., & Grompe, M. (2016). A universal system to select gene-modified hepatocytes in vivo. Science Translational Medicine, 8(342), [342ra79]. https://doi.org/10.1126/scitranslmed.aad8166

A universal system to select gene-modified hepatocytes in vivo. / Nygaard, Sean; Barzel, Adi; Haft, Annelise; Major, Angela; Finegold, Milton; Kay, Mark A.; Grompe, Markus.

In: Science Translational Medicine, Vol. 8, No. 342, 342ra79, 08.06.2016.

Research output: Contribution to journalArticle

Nygaard, S, Barzel, A, Haft, A, Major, A, Finegold, M, Kay, MA & Grompe, M 2016, 'A universal system to select gene-modified hepatocytes in vivo', Science Translational Medicine, vol. 8, no. 342, 342ra79. https://doi.org/10.1126/scitranslmed.aad8166
Nygaard S, Barzel A, Haft A, Major A, Finegold M, Kay MA et al. A universal system to select gene-modified hepatocytes in vivo. Science Translational Medicine. 2016 Jun 8;8(342). 342ra79. https://doi.org/10.1126/scitranslmed.aad8166
Nygaard, Sean ; Barzel, Adi ; Haft, Annelise ; Major, Angela ; Finegold, Milton ; Kay, Mark A. ; Grompe, Markus. / A universal system to select gene-modified hepatocytes in vivo. In: Science Translational Medicine. 2016 ; Vol. 8, No. 342.
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