MicroRNAs regulatory networks in cardiotoxicity

A long time and many steps are needed to bring valuable drugs to patients. Drugs not only have to be effective against the disease which they are intended for, but also safe. All drugs cause toxic effects in proportion to the dose (Paracelsus, ca. XVI cent.). Therefore, assessing carefully the optimal dose at the desired benefit/risk ratio (Stevens and Baker, 2009) and characterizing mechanisms of toxicity are essential for maintaining high safety standards. The experimental strategies used to predict drugs’ adverse events are many, and include the use of non-invasive biomarkers that can ideally be translated from test species to humans. In particular, cardiac safety requires special attentions, as it causes the cessation of drug development in over the 30% of cases due to toxic liabilities in Phase III, and over 40% post-approval (Redfern et al., 2011). In 1993 Victor Ambros, Rosalind Lee and Rhonda Feinbaum (Lee et al., 1993) described for the first time in C. elegans the role of short non-coding RNAs, called microRNAs. MicroRNAs pair with target mRNAs via sequence complementarity, and lead to mRNA degradation or translation inhibition (Filipowicz et al., 2008; Guo et al., 2010). MicroRNAs are implicated in many biological processes and in the evolution of the complexity of superior organisms, for review see (Bartel, 2004; Berezikov, 2011). Their role in cardiovascular development and diseases has been supported by an increasing number of publications, for review see (Kinet et al., 2012; Small and Olson, 2011). Recently, tissue-specific microRNAs were found in a variety of body fluids following drug-induced tissue injury (Wang et al., 2009). In particular, myocardial necrosis of different etiologies (such as acute coronary syndrome and myocardial infarction) caused an increase of heart enriched microRNAs in plasma/serum of patients, as reviewed by (McManus and Ambros, 2011). This work is aimed at increasing our knowledge about cardiac microRNAs function in different toxicological contexts and across species. Chapter 1 will give an overview of drug safety assessment focusing on the cardiovascular system, microRNAs biology and their potential as toxicity biomarkers. In Chapter 2 I will illustrate an example of drug-induced cardiac injury, and its impact on expression of microRNAs in rats’ hearts. Chronic treatment of rats with low amounts of doxorubicin (an anti-tumoral compound) caused myocardial vacuolation in ventricular tissue and a significant increase of specific microRNAs and genomic indicators of cardiomyopathy. We found that Sipa1 could be directly inhibited by miR-34c by using a luciferase assay. Notably, the phenotypic anchoring of microRNAs with histopathological read-outs in the tissue showed that over-expression of miR-216b in the heart preceded the rise of overt lesions. Chapter 3 will describe an approach to determine the distribution of mRNAs and microRNAs in different cardiac structures in rat, dog and Cynomolgus monkey. We showed that microRNAs could discriminate the different heart structures as good as mRNAs. We also assessed the expected anti-correlation between microRNAs levels and their predicted target mRNAs, and showed that mRNAs of 4 genes implicated in cardiac diseases and muscular development could be inhibited at the post-transcriptional level by 4 distinct microRNAs. We propose that our mRNAs and microRNAs data sets could be used to assess the human relevance of preclinical findings in these 3 species, and to derive potential tissue and circulating biomarkers for drug-induced cardiac injury. In conclusion, profiling microRNAs have the potential to help biomedical research in bringing drugs to patients, since they can shed light on toxicity mechanisms in the tissues (Taylor and Gant, 2008) and are a source of translatable non-invasive toxicity biomarkers in body fluids.