TOP-EVs: Technology Of Protein delivery through Extracellular Vesicles to target PCSK9

High total cholesterol levels are a major cause of disease burden in both the developed and developing world as a risk factor for ischemic heart disease and stroke. Lowering cholesterol has been one of the most effective strategies to prevent major cardiovascular events. During the life-long lowering cholesterol management, many patients experience intolerant side-effect, or develop CVD despite being on maximally dose therapy, or being non-responsive hypercholesterolemia or too expensive to afford, thus there is an unmet need for novel long-lasting therapy. Recent developed biotechnologies provide an alternative strategy for lowering cholesterol levels: permanent removal of

PCSK9, a hepatic protease that cause cholesterol accumulation indirectly. In this project, we established a technology that allows a future new treatment for hypercholesterolemia by permanently removing PCSK9 from hepatocytes by using cell-derived nanomedicines, called extracellular vesicles.

In Technology Of Protein delivery through Extracellular Vesicles (TOP-EV), we enabled protein loading inside Extracellar vesicles (EVs), allowing a protein of interest to be delivered to target cells. The goal is to employ the TOP-EVs to delivery CRISPR/Cas9 to the liver and thereby targeting PCSK9. We established the loading of the different proteins, including CRIPSP/Cas9 and induced a permanent loss of PCSK9 in human and mouse liver cells. We managed to upscale the production of this product and prepared large batches of the TOP-EVs to demonstrate feasibility of PCSK9 removal in animals.

At the end of the project, we were able to show the feasibility of our innovative EV-loading strategy and the first successful in vivo results of this approach. Currently, we are securing protection of our inventions, starting a new private environment and make the first steps to confirm the long-lasting genetic removal in vivo.