Bile-duct proliferation as an unexpected side-effect after AAV2-LDLR gene transfer to rabbit liver

Elisa Hytönen,Nihay Laham-Karam,Mika Hujo,Hanna Kankkonen,Anniina Laurema,Atsushi Miyanohara,Seppo Ylä-Herttuala,Vesa Kärjä

doi:10.1038/s41598-019-43459-1

Abstract

Familial hypercholesterolemia (FH) is an inherited disease of lipoprotein metabolism caused by a defect in the LDL receptor (LDLR) leading to severe hypercholesterolemia, and associated with an increased risk of coronary heart disease and myocardial infarction. We have developed a gene therapy protocol for FH using AAV2, AAV9 and lentiviral vectors and tested safety and efficacy in LDL receptor deficient Watanabe Heritable Hyperlipidemic rabbits. We show that LV-LDLR produced a significant long-lasting decrease in total serum cholesterol whereas AAV9-LDLR resulted only in a transient decrease and AAV2-LDLR failed to reduce serum cholesterol levels. A significant pathological side effect, bile-duct proliferation, was seen in the liver of AAV2-LDLR rabbits associated with an increased expression of Cyr61 matricellular protein. Special attention should be given to liver changes in gene therapy applications when genes affecting cholesterol and lipoprotein metabolism are used for therapy.

Highlights

Familial hypercholesterolemia (FH) is an inherited disease of lipoprotein metabolism caused by a defect in the low-density lipoprotein (LDL)-receptor (LDLR), leading to reduced turnover of LDL and significantly increased plasma cholesterol levels
We have previously shown that lentivirus (LV) -mediated LDL receptor (LDLR) gene transfer to Watanabe heritable hyperlipidemic (WHHL) rabbit liver led to a long-term significant decrease in serum cholesterol levels[7]
The decline in total cholesterol levels four weeks after LV-LDLR gene transfer was on average 18 ± 8% and cholesterol values continued to decrease to 47 ± 9% below pre-treatment values one year after gene transfer compared to an increase of 6 ± 15% in LV-control animals (p = 0.033)

Summary

Introduction

Familial hypercholesterolemia (FH) is an inherited disease of lipoprotein metabolism caused by a defect in the LDL-receptor (LDLR), leading to reduced turnover of LDL and significantly increased plasma cholesterol levels. We have previously shown that lentivirus (LV) -mediated LDLR gene transfer to WHHL rabbit liver led to a long-term significant decrease in serum cholesterol levels[7] We extended these studies to a full preclinical safety and toxicology study in WHHL rabbits, and explored the potential of another long-term vector, AAV, in the treatment of FH. We describe a surprising new finding of bile duct proliferation associated with increased Cysteine Rich Angiogenic Inducer 61 (Cyr61) expression in the liver after AAV2-mediated liver-directed gene transfer To our knowledge, this is the first study to compare these vector systems for the treatment of FH in a preclinical safety, efficacy and toxicology study to select the optimal vector for further clinical development

Methods

Results

Conclusion