Genome editing of fructose metabolism to correct hereditary fructose intolerance in mice.

About This Grant

PROJECT SUMMARY/ABSTRACT. Excessive intake of sugar, and particularly fructose, has been linked to different metabolic health issues, including obesity, diabetes, and cardiovascular disease. Part of the reason for the strong association between high fructose consumption with metabolic diseases is because fructose -as sugar- is often added to processed foods to increase their palatability. One group that is particularly sensitive to the deleterious metabolic consequences of fructose is people with hereditary fructose intolerance (HFI). HFI is a genetic disease triggered by fructose that is caused by mutations in the aldob gene with serious health implications. As a result, individuals with HFI have defective fructose metabolism and accumulate this sugar as fructose-1-phosphate (F1P). This leads to severe reactions following fructose ingestion such as abdominal pain, symptomatic hypoglycemia, lactic acidosis, seizures and chronic metabolic consequences including growth retardation, chronic kidney disease and metabolic dysfunction-associated steatohepatitis. Further, if the exposure to fructose is continuous, HFI can be fatal. Unfortunately, there is no cure for HFI and therefore, any potential therapeutical approach needs to be explored. In this application, we continue the work from the previous award to investigate molecular mechanisms and metabolic pathways that we can target to treat this condition. We have identified that targeting ketohexokinase, KHK, the enzyme located upstream AldoB in the metabolic pathway of fructose, is a promising strategy to treat HFI as it will prevent the formation and accumulation of F1P. This finding has recently been confirmed in a proof- of-concept study in three individuals with HFI receiving a specific KHK inhibitor. However, further studies are necessary to assess individual safety with longer follow-up and clinically relevant endpoints. Here, we have developed a novel murine model of HFI, the aldobA149P mouse- which harbors an ortholog mutation of the most frequent variant in human HFI (the A149P). Initial characterization of aldobA149P mice demonstrate that this strain phenocopies the human disease and develop clear aversion to fructose. With this model, we propose to study two different yet complementary strategies to treat HFI. In the first aim of the application, we propose to conditionally repress KHK expression in selected cell types -enterocytes and hepatocytes- in adult aldobA149P mice and evaluate the subsequent behavioral and metabolic response to fructose. We will also evaluate if targeting KHK is a promising approach to reverse chronic HFI in mice. Second, we propose to perform CRISPR/Cas9-mediated in vivo gene editing to repair and correct the A149P missense mutation in the aldob gene of adult aldobA149P mice. Then, we will induce clonal selection and repopulation of the liver with repaired hepatocytes by exposing mice to fructose after which, fructose tolerability, the metabolic response to fructose and analysis of potential in vivo off-target effects will be evaluated.