Amsterdam University Medical Centers, University of Amsterdam, Tytgat Institute for Liver and Intestinal Research, Amsterdam Gastroenterology and Metabolism, Meibergdreef 9, Amsterdam, The Netherlands.
Amsterdam University Medical Centers, University of Amsterdam, Department of Pathology, Meibergdreef 9, Amsterdam, The Netherlands.
J Hepatol. 2019 Jul;71(1):153-162. doi: 10.1016/j.jhep.2019.03.021. Epub 2019 Mar 29.
BACKGROUND & AIMS: Progressive familial intrahepatic cholestasis type 3 (PFIC3), for which there are limited therapeutic options, often leads to end-stage liver disease before adulthood due to impaired ABCB4-dependent phospholipid transport to bile. Using adeno-associated virus serotype 8 (AAV8)-mediated gene therapy, we aimed to restore the phospholipid content in bile to levels that prevent liver damage, thereby enabling stable hepatic ABCB4 expression and long-term correction of the phenotype in a murine model of PFIC3.
Ten-week-old Abcb4 mice received a single dose of AAV8-hABCB4 (n = 10) or AAV8-GFP (n = 7) under control of a liver specific promoter via tail vein injection. Animals were sacrificed either 10 or 26 weeks after vector administration to assess transgene persistence, after being challenged with a 0.1% cholate diet for 2 weeks. Periodic evaluation of plasma cholestatic markers was performed and bile duct cannulation enabled analysis of biliary phospholipids. Liver fibrosis and the Ki67 proliferation index were assessed by immunohistochemistry.
Stable transgene expression was achieved in all animals that received AAV8-hABCB4 up to 26 weeks after administration. AAV8-hABCB4 expression restored biliary phospholipid excretion, increasing the phospholipid and cholesterol content in bile to levels that ameliorate liver damage. This resulted in normalization of the plasma cholestatic markers, alkaline phosphatase and bilirubin. In addition, AAV8-hABCB4 prevented progressive liver fibrosis and reduced hepatocyte proliferation for the duration of the study.
Liver-directed gene therapy provides stable hepatic ABCB4 expression and long-term correction of the phenotype in a murine model of PFIC3. Translational studies that verify the clinical feasibility of this approach are warranted.
Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a severe genetic liver disease that results from impaired transport of lipids to bile, which makes the bile toxic to liver cells. Because therapeutic options are currently limited, this study aims to evaluate gene therapy to correct the underlying genetic defect in a mouse model of this disease. By introducing a functional copy of the missing gene in liver cells of mice, we were able to restore lipid transport to bile and strongly reduce damage to the liver. The proliferation of liver cells was also reduced, which contributes to long-term correction of the phenotype. Further studies are required to evaluate whether this approach can be applied to patients with PFIC3.
进行性家族性肝内胆汁淤积症 3 型（PFIC3）的治疗选择有限，由于 ABCB4 依赖的磷脂向胆汁的转运受损，常导致患者在成年前发展为终末期肝病。我们采用腺相关病毒血清型 8（AAV8）介导的基因治疗，旨在将胆汁中的磷脂含量恢复到可防止肝损伤的水平，从而使 PFIC3 小鼠模型中稳定表达 ABCB4 并长期纠正表型。
10 周龄 Abcb4 小鼠通过尾静脉注射，接受单次剂量的 AAV8-hABCB4（n=10）或 AAV8-GFP（n=7），均受肝脏特异性启动子控制。载体给药后 10 或 26 周时，动物被处死，以评估转基因的持久性，并在接受 0.1%胆酸盐饮食 2 周后进行挑战。定期评估血浆胆汁淤积标志物，并进行胆管插管以分析胆汁磷脂。通过免疫组化评估肝纤维化和 Ki67 增殖指数。
所有接受 AAV8-hABCB4 治疗的动物在给药后 26 周内均实现了稳定的转基因表达。AAV8-hABCB4 的表达恢复了胆汁磷脂的排泄，将胆汁中的磷脂和胆固醇含量增加到可改善肝损伤的水平。这导致血浆胆汁淤积标志物碱性磷酸酶和胆红素恢复正常。此外，AAV8-hABCB4 还可预防进行性肝纤维化，并在研究期间降低肝细胞增殖。
肝靶向基因治疗为 PFIC3 小鼠模型提供了稳定的肝 ABCB4 表达和长期的表型纠正。需要进行验证该方法临床可行性的转化研究。
进行性家族性肝内胆汁淤积症 3 型（PFIC3）是一种严重的遗传性肝脏疾病，由脂质向胆汁的转运受损引起，使胆汁对肝细胞有毒。由于目前治疗选择有限，本研究旨在评估基因治疗在该疾病的小鼠模型中纠正潜在的遗传缺陷。通过在小鼠肝细胞中引入缺失基因的功能性副本，我们能够恢复脂质向胆汁的转运，并大大减少对肝脏的损伤。肝细胞的增殖也减少了，这有助于长期纠正表型。需要进一步研究以评估该方法是否可应用于 PFIC3 患者。
