Within LONGHEART we focused on the large fraction of long non-coding RNAs (lncRNAs) and their functional roles, as well as diagnostic and therapeutic use in heart failure. The proposal had the following interconnected objectives and I will briefly describe what we have done here in the 5-6 years:
1: identify novel functional relevant cardiac remodeling-associated lncRNAs;
We have implemented novel high-throughput methods for lncRNA identification and identified several lncRNA candidates as interesting novel therapeutic targets. For example lncRNA Chast (cardiac hypertrophy-associated transcript) (Science Transl Med. 2016;8:326ra22.) which triggers pathological growth and further remodelling processes in the heart, which ultimately lead to weakening of the heart function. Silencing of Chast both prevented and attenuated TAC (transverse aortic constriction)-induced pathological cardiac remodeling with no early signs on toxicological side effects. The lncRNA Meg3 (maternally expressed gene 3) was found to be a novel and innovative target in heart failure pathologies with fibrosis (Circ Res. 2017;121:575-583.). Besides the identification of several other lncRNAs we created a lentiviral shRNA library-based approach for functional lncRNA profiling (Cell Death Differ. 2018;25:307-318.). With this study we were able to demonstrate the applicability of an innovative shRNA library approach to identify long non-coding RNA functions in a massive parallel approach.
2: Characterisation of key lncRNA cardiac targetomes
Here, we showed that Chast interacts mainly with proteins involved in cardiomyopathies and correlates with cardiac disease pathways, qualifying Chast as a potential target for therapeutics (Science Transl Med. 2016;8:326ra22.). In a further study, it was shown that the lncRNA H19 is down-regulated in failing hearts from mice, pigs and humans (Eur Heart J, 2020 Sep 21;41(36):3462-3474). H19 acts as an anti-hypertrophic lncRNA and therefore represents a promising therapeutic target to combat pathological cardiac remodelling.
3: Investigation of lncRNA-paracrine mechanisms and the diagnostic and prognostic potential of cardiac-derived extracellular lncRNAs using large clinical cohorts
We discovered lncRNAs secreted by hypoxic cardiomyocytes. Specifically, we have unraveled a novel intercellular communication route between hypoxic cardiomyocytes and fibroblasts via extracellular vesicles, which are enriched with lncRNAs. Vesicles were taken up by fibroblasts, triggering expression of profibrotic genes. Genetic loss of a certain lncRNA in vivo resulted in an impaired heart function after myocardial infarction highlighting its translational relevance (Mol Ther Nucleic Acids, 2019;18:363-374).
4: Discover the therapeutic potential to prevent cardiac remodeling in clinically relevant animal models.
We have developed cell type specific targeting vectors for the modulation of cardiac lncRNAs and have provided first evidence for therapeutic manipulation of lncRNAs in heart diseases (Science Transl Med. 2016;8:326ra22). We could show that in vivo treatment of mice after pressure overload of the left ventricle develop significantly less cardiac fibrosis and have improved diastolic function. (Circ Res. 2017 Aug 18;121(5):575-583). Finally, we reported on the powerful therapeutic potential of the conserved lncRNA H19 in the treatment of pathological cardiac hypertrophy. Pressure overload-induced cardiac hypertrophy in H19 knock-out mice was aggravated compared to wild-type mice. In contrast, vector-based, cardiomyocyte-directed gene therapy using murine and human H19 strongly attenuated heart failure even when cardiac hypertrophy was already established. H19 was highly conserved and down-regulated in failing hearts from mice, pigs and humans. We showed H19 gene therapy to prevent and reverse experimental pressure-overload-induced heart failure (Eur Heart J, 2020 ;41:3462-3474.).
Within the framework of the funding, many promising lncRNAs have been identified and functionally characterised and analyses have also been carried out with regard to their diagnostic and therapeutic potential. These were the objectives of the project and this was fully achieved.
