Molecular Genetic Approach to the Study of Familial Dilated Cardiomyopathies

In this thesis, I will report on the results of some molecular genetic studies carried out on patients with three inherited forms of Dilated Cardiomyopathy (DCM). DCM is defined as a primary disease of the myocardium associated with dilatation and impaired contraction of the left ventricle or both ventricles. Dilated cardiomyopathy represents a major cause of heart failure, arrhythmias, morbidity and mortality among cardiovascular diseases and it is the leading indication for heart transplantation. The occurrence of a familial form of dilated cardiomyopathy was very rarely reported in the past, but recent studies revealed a frequency of at least 20-25% of genetic transmission. Different modes of inheritance were recognized for the familial form: an autosomal dominant form, which is the most frequent form of familial dilated cardiomyopathy (AD-FDC); an autosomal recessive; an X-linked (XLDC), an autosomal dominant form with conduction defects and later development of DC (CDDC), and finally a newly recognized autosomal dominant form with clinical signs of myopathy (MDDC). The population study was carried out on a total of 60 families for a total number of 255 evaluated individuals. Our attention was focused on three of these different forms of DCM. Two families with suggestive X-linked mode of inheritance underwent linkage analysis which confirmed the association of the disease with the dystrophin gene. Further molecular analysis at the DMD gene level, allowed us to identify in one family a point mutation at the first muscular exonintron boundary, abolishing the splicing of the muscular specific mRNA. In the second family, similarly to what occurs to Becker Muscular Dystrophy patients, a deletion of exons 48-49 was found by multiplex PCR analysis. Among the different forms of DCM there is a newly identified form of FDC which shows additional signs of myopathy (MDDC). Since MDDC was recalling the better known XLDC form, determined by mutations of dystrophin, a candidate genes linkage analysis was performed for more than 30 loci where candidate genes coding for structural proteins of the cytoskeleton, the sarcolemma or the extra cellular matrix are localized. Unfortunately, this approach turned out to be unsuccessful. The most frequent form of DCM is the autosomal dominant form of FDC (AD-FDC) which occurs in 55% of cases. Using a genome-scan approach, a first locus for AD-FDC was mapped in a large family pedigree (> 120 examined members) and confirmed in other two kindreds with AD-FDC. A cumulative lod score value of 3.69 was obtained with the polymorphic marker D9S153. Subsequently, the candidate genes which map within the identified interval were analyzed. However, none was found in linkage with FDC. Since the candidate gene approach was unsuccessful, the map refinement of the region was carried out using new genetic markers which became available in the data banks. This map refinement led to the detection of new recombination events in critical patients, which dropped down the informativity of the identified FDC locus. Similarly, also analysis of the D9S156-D9S157 interval in chromosome 9, another locus with suggestive linkage at the first genome scan, did not produce further evidence for linkage. These unconclusive results underline the problems that still hamper worldwide the identification of the genes responsible for FDC. These problems are related to the phenotypic complexity of the disease, its low penetrance, its genetic heterogeneity and the high chance of misleading clinical status definition. All these variables greatly limit the efficacy of a linkage analysis approach for this disease.