Mendelian Genetics of Stroke: an assessment of the broad genetic and phenotypic heterogeneity driven by the identification of underlying pathogenic mechanisms

Being the second cause of death and disability world-wide, approximately 1.1 million inhabitants of Europe suffer a stroke each year, the ischemic stroke accounting for approximately 80% of cases. Stroke presents different prevalence for the diverse range of age: perinatal stroke (1 per 2700), pediatric stroke (1-2 in 100000), juvenile stroke (1 per 10000), adult stroke (95-290 in 100000). Evidence for a genetic aetiology of stroke has largely been described in the literature. Juvenile and pediatric stroke, more seldom neonatal and adult stroke, is thought to be mainly driven by different underlying genetic mechanisms: several mendelian disorders are known to be the cause of monogenic form of stroke. Next Generation Sequencing (NGS) panels represent an excellent option for diagnostic routine. Genotyping of stroke patients is fundamental for focused clinical follow-up and family counselling. Albeit NGS-panels represent an excellent option for diagnostic routine, the mutation detection rate turns out to be ≈10%, even for patients affected with a likely monogenic stroke. Whole exome sequencing (WES) embodies straightforward approach in this context, allowing the identification of novel candidate genes. The present project has focused on: i) the identification of the underlying genetic causes of monogenic stroke cases, ii) the improvement of our knowledge about clinically overlapping phenotypes through the genetic characterization of the corresponding patients, iii) the optimization of the diagnostic work-up in order to administer disease specific follow-up and treatments to patients. A custom NGS-panel including 183 genes associated with monogenic forms of stroke was designed according to clinicians and neuroradiologists belonging to the multidisciplinary team of Gaslini Stroke Center. Patients affected by suspected monogenic stroke were selected by different Unit of Gaslini Institute and other Italian children/adult hospitals collaborating with the Gaslini Stroke Center. DNA samples of 102 individuals were extracted and analyzed via the mentioned multigenic panel. Specific functional assays were set up and optimized in order to dissect molecular disease mechanisms in selected subgroup or single patients. Moreover, WES has been applied in specific trios after a negative result of the panel. Up to 16 of 102 harbored pathogenic variants justifying or potentially justifying their clinical and neuroradiological phenotypes. The overall mutational rate is around 16.5%. Specific functional assays opened the way to understand molecular disease mechanism underlying different type of monogenic stroke. As the number of genes involved in the stroke pathogenesis is rapidly growing, a WES approach coupled with an in-silico gene panel has more recently been applied. This not only allow to test a constantly updated gene list but also to look at the rest of the genes of the genome in case the in-silico panel resulted negative. In conclusion, the project of the thesis allowed to identify causative mutation in some of the patients suspected to have a monogenic stroke and to dissect potential molecular mechanisms underlying their stroke subtype. This let the establishment of a more specific follow-up and counselling for families feasible.