Article ; Online: Resolution of hepatic fibrosis after ZFN-mediated gene editing in the PiZ mouse model of human α1-antitrypsin deficiency.
Hepatology communications
2023 Volume 7, Issue 3, Page(s) e0070
Abstract: Background: α1-antitrypsin deficiency is most commonly caused by a mutation in exon-7 of SERPINA1 (SA1-ATZ), resulting in hepatocellular accumulation of a misfolded variant (ATZ). Human SA1-ATZ-transgenic (PiZ) mice exhibit hepatocellular ATZ ... ...
| Abstract | Background: α1-antitrypsin deficiency is most commonly caused by a mutation in exon-7 of SERPINA1 (SA1-ATZ), resulting in hepatocellular accumulation of a misfolded variant (ATZ). Human SA1-ATZ-transgenic (PiZ) mice exhibit hepatocellular ATZ accumulation and liver fibrosis. We hypothesized that disrupting the SA1-ATZ transgene in PiZ mice by in vivo genome editing would confer a proliferative advantage to the genome-edited hepatocytes, enabling them to repopulate the liver. Methods: To create a targeted DNA break in exon-7 of the SA1-ATZ transgene, we generated 2 recombinant adeno-associated viruses (rAAV) expressing a zinc-finger nuclease pair (rAAV-ZFN), and another rAAV for gene correction by targeted insertion (rAAV-TI). PiZ mice were injected i.v. with rAAV-TI alone or the rAAV-ZFNs at a low (7.5×1010vg/mouse, LD) or a high dose (1.5×1011vg/mouse, HD), with or without rAAV-TI. Two weeks and 6 months after treatment, livers were harvested for molecular, histological, and biochemical analyses. Results: Two weeks after treatment, deep sequencing of the hepatic SA1-ATZ transgene pool showed 6%±3% or 15%±4% nonhomologous end joining in mice receiving LD or HD rAAV-ZFN, respectively, which increased to 36%±12% and 36%±12%, respectively, 6 months after treatment. Two weeks postinjection of rAAV-TI with LD or HD of rAAV-ZFN, repair by targeted insertion occurred in 0.10%±0.09% and 0.25%±0.14% of SA1-ATZ transgenes, respectively, which increased to 5.2%±5.0% and 33%±13%, respectively, 6 months after treatment. Six months after rAAV-ZFN administration, there was a marked clearance of ATZ globules from hepatocytes, and resolution of liver fibrosis, along with reduction of hepatic TAZ/WWTR1, hedgehog ligands, Gli2, a TIMP, and collagen content. Conclusions: ZFN-mediated SA1-ATZ transgene disruption provides a proliferative advantage to ATZ-depleted hepatocytes, enabling them to repopulate the liver and reverse hepatic fibrosis. |
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| MeSH term(s) | Humans ; Animals ; Mice ; Gene Editing ; Zinc Finger Nucleases ; Liver Cirrhosis/genetics ; Liver Cirrhosis/therapy ; Hepatocytes ; Disease Models, Animal ; Intracellular Signaling Peptides and Proteins |
| Chemical Substances | Zinc Finger Nucleases (EC 3.1.-) ; Intracellular Signaling Peptides and Proteins |
| Language | English |
| Publishing date | 2023-02-27 |
| Publishing country | United States |
| Document type | Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't |
| ISSN | 2471-254X |
| ISSN (online) | 2471-254X |
| DOI | 10.1097/HC9.0000000000000070 |
| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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