Toll-like receptor 3 mediates Calcific Aortic Valve Disease

Background: Calcific aortic valve disease (CAVD) disease is the third leading cause of cardiovascular related disease and represents therefore a major socioeconomic burden in the western world. Currently, the only treatment option is aortic valve replacement. CAVD is caused by an osteoblastic phenotype switch of valvular cells. However, the trigger for the phenotype switch remains unknown. Aims: Toll-like receptor 3 (TLR3) is part of the innate immune system activated by viral and endogenous RNA released from dying cells. We hypothesized that mechanical strain leads to TLR3 activation leading to an osteoblastic phenotype switch of valvular cells with subsequent initiation of calcific aortic valve disease. This project aims to (a) clarify the role of TLR3 in the pathogenesis of CAVD and (b) investigate TLR3 as potential therapeutic target for the prevention of CAVD development. Methods: Aortic valves are obtained from patients undergoing aortic valve replacement or from explanted hearts. Valvular cells are isolated and treated with receptor agonists or antagonists. Osteoblastic gene expression is evaluated. Cells are challenged with osteoblastic medium and analyzed for calcific activity. A specific system is used to apply mechanical strain to valvular cells. Aortic valve morphology and function of specific knock out mice lacking TLR3 is analyzed via transthoracic echocardiography, microCT and histological evaluation. Innovation of the project: Currently, there is no therapeutic treatment option for the prevention of CAVD. The proposed project offers a straight-forward approach to test the hypothesis in (a) cell culture experiments with primary aortic valve cells, (b) extensive morphological and functional characterization of aortic valves in animal models and (c) analysis of patient material. Preliminary results show promising results: Mice lacking TLR3 do not develop CAVD. TLR3 could become an effective target for the pharmacological prevention of CAVD. A pharmacological treatment option preventing the progression of the disease could spare many patients from invasive open heart surgeries.

Project Summary (English) Targeting inflammation to stop heart valve calcification: a new path toward drug therapy Calcification of the aortic heart valve, known as calcific aortic valve disease (CAVD), is one of the most common heart conditions in aging populations. Despite its rising impact, there are still no medications available to prevent or slow this disease - currently, heart valve replacement through surgery or catheter-based interventions is the only effective treatment. Our research has identified a key mechanism behind valve calcification: a specific immune receptor called Toll-like receptor 3 (TLR3) becomes activated, not only during infections but also by stress signals from within damaged heart tissue. This activation triggers inflammation and causes cells in the heart valve to change their identity - becoming more like bone-forming cells, which leads to calcification of the valve. In our project, we discovered that inhibiting the TLR3 pathway can effectively block this harmful process. We tested several existing drugs and a new type of RNA-based therapy called antisense oligonucleotides (ASOs) that can precisely block one of the key proteins in this pathway, IRF3. Both strategies showed striking effects: they prevented calcification in laboratory cell models and in mice, without causing significant side effects. This is a major step forward in identifying a first-in-class therapeutic approach to treat or even prevent aortic valve calcification. Our work suggests that repurposing existing safe medications or developing targeted RNA therapies could offer future alternatives to surgery . Moreover, the screening methods we developed could be applied to other inflammatory diseases with similar calcification mechanisms. This research was made possible through support from the Austrian Science Fund (FWF) and marks an important contribution from Austria to the global effort of developing non-invasive treatments for heart valve disease.