Applicability, efficacy and safety of targeted vehicle mediated in vivo base editing in a mouse model of hereditary hemochromatosis type I

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Hemochromatosis is one of the most common inherited metabolic diseases among white populations and predominantly originates from a homozygous C282Y mutation in the HFE gene. The G > A transition at position c.845 of the gene causes misfolding of the HFE protein, ultimately resulting in its absence at the cell membrane.This consequently leads to continuous iron uptake in the intestine and accumulation of iron in various organs such as liver, pancreas and heart. Affected patients may exhibit liver cirrhosis, diabetes mellitus as well as cardiomyopathy. The current state-of-the-art treatment of hemochromatosis includes life-long phlebotomy and the application of chelating agents such as deferoxamine. In a mouse model (129-HFE tm 1.1Nca), which carries the human mutation, we aim to correct the mutation in vivo by application of a targeted ABE base editor RNA. In this proposal we will develop advanced transient ABE base editor therapies either based on mRNA/gRNA molecules incorporated into lipid nanoparticles (LNP) or into virus-like particles (VLP) to correct the C282Y point mutation. Initially, we will perform dose finding and toxicity studies by analyzing liver enzymes and cytokine levels. Then, we will perform long-term experiments for up to 12 months to demonstrate efficacy and genotoxicity. Efficacy of the therapeutic approach will be analysed by the rate of base conversion as measured by DNA sequencing and the normalisation of hemochromatosis relevant blood parameters. Genotoxicity of the base editor therapy will analysed on a global scale by searching for alterations at DNA and RNA levels, which could occur in other regions than the target sequence (e.g. "off targets"). Overall, our experiments will provide the basis for a safe and efficient therapy for hereditary hemochromatosis and a blueprint for the treatment of other hereditary liver diseases.