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Cardiovascular diseases (CVD) and their associated pathologies are among the greatest causes of morbidity and mortality, entailing approximately 17.3 deaths a year.1 This class of pathology as a whole has a multi-factor etiology. Its possible prognoses lead to public health issues, with its incidence being related to behavioral, metabolic and genetic risk factors.2 In spite of the fact that the treatments established for the CVDs and their possible prognoses decrease the rhythm of progression of the illness, the need to develop therapeutic approaches able to reverse the pathology and its complications is growing.  The progresses in the fields of molecular and cellular biology have allowed the elucidation of molecular pathways and genetic causes involved in the establishment and progression of the CVDs, outlining a new viewpoint with regard to the prevention, treatment and possible outcomes of this pathological class. Recent discoveries, both experimental and those obtained by means of bioinformatics tools, regarding the molecular bases of cardiovascular dysfunctions, have been pointing to considerable therapeutic targets.3 However, most of these targets cannot be pharmacologically manipulated, which makes them potential candidates for genic therapy, such as the factors involved, for example, in angiogenesis, apoptosis and endothelial dysfunction.4 Within such context, gene manipulation may help suppress genetic factors connected to the incidence of the CVDs, as well as to mitigate the clinical complications caused by ischemic and occlusive events. Thus, the development and improvement of genome edition tools allow the creation of therapies focused in the genetic risk factors to cardiovascular damage and fundamental morphophysiological issues caused by the CVDs. In such context, the system formed by clustered regularly interspaced short palindromic repeats (CRISPR), and its CRISPR associated protein-9 (Cas9), stands out due to how easy it is to use it, its high specificity, easy in vitro and in vivo manipulation, in addition to the possibility of simultaneously editing multiple targets. Given the genomic complexity that intervenes in the CVDs, we shall indicate herein certain possibilities of applying the CRISPR/Cas9 tool in Cardiology.  Unraveling the CRISPR/Cas9 system Developed from molecular organisms of the bacterial immune system, the CRISPR system allows the edition of the genome by means of splicing of the DNA by an endonuclease (Cas9), guided based on an RNA sequence, which is able to pair up with the bases of a target sequence (Figure 1).5 The CRISPR genetic structure, in the bacterial system, is made up of clustered regularly interspaced short palindromic repeats. The repeats and the spacers (which may contain interspacing viral sequences), when transcribed, form the transactivator RNA (or guide RNA), which serves to direct the Cas9 enzyme, a nuclease, to the target (in this case, the parasite virus sequence). Taking advantage of this strategy, both the Cas9 protein and the guide RNA can be introduced in vitro into other cells and directed to specific places in the genome, for them to cause breaks to the double strand. After this splicing, the intrinsic molecular machinery of the organism, responsible for the correction of errors in the genome, is used to alter the DNA sequence, adopting the modification. The system can thus be used both to repair mutations (restoring genic function) and to introduce new mutations (causing the genic "knockout"). Therefore, by conciliating sophisticated molecular and biotechnological techniques, the CRISPR/Cas9 system was proposed for application on genomic editing and is currently commercially available for thousands of targets.6 Both the RNA and the Cas9 protein, produced in vitro, can be delivered to the cells using different mechanisms, such as the use of vectors or chemical agents.    Figure 1  CRISPR/Cas9 system - target recognition mechanism. The guide RNA is designed to recognize the target sequence to be modified in the RNA and introduce modifications. When the pairing of nitrogenized bases occurs (due to the annealing of the target sequence ...      The most simple applicability of the CRISPR system is connected to changing single or certain bases in genes with a well-defined allelic relationship. It is important to stress that this relationship of Mendelian dominance must be taken into account for the genic function to be achieved, both to activate it and to inhibit it. However, bi-allelic modifications have also been successfully obtained7. Moreover, the use of the CRISPR/Cas9 has also been proposed for embryonic stage in animal models, where the progeny can generate "founding" organisms (by recombination) containing allelic mutations that lead to the "knockout" effect or with diminished expression.8 In such context, the CRISPR/Cas9 system is being quickly adopted to edit and modify genomes in several cellular types, including stem cells,9 and has been giving good results in the edition of human genes.10 The press recently reported that researchers from the University of Pennsylvania have been given approval from the Food and Drug Administration (FDA) to conduct a clinical study to begin in 2017, the targets of which are 3 genes involved in cancer. Thus, the question arises regarding the possibility of applying the CRISPR/Cas9 system to such a biologically complex situation as cardiovascular diseases.

The CRISPR/Cas9 System and the Possibility of Genomic Edition for Cardiology