Utvrđivanje genetičke osnove retkih neurodegenerativnih bolesti analizom kliničkog egzoma

Abstract

eurodegenerativne bolesti se odlikuju varijabilnom kliničkom slikom, vremenom početka bolesti, prognozom i odgovorom na lečenje. Najveći broj ovih oboljenja uključuje motorne i kognitivne poremećaje poput Alchajmerove i Parkinsonove bolesti koje su široko rasprostranjene, do izuzetno retkih poput Krojcfeld-Jakobove bolesti. Ovom doktorskom tezom istražena je genetička osnova kognitivnih poremećaja i poremećaja pokreta sekvenciranjem panela klinički egzom kod ukupno 57 nesrodnih bolesnika (15 sa kognitivnim poremećajem i 42 sa poremećajem pokreta). Prednost pri selekciji su imali porodični slučajevi sa ranim početkom bolesti ili složenim fenotipom. Sekvenciranje panela vršeno je na Illumina MiSeq platformi prema uputstvu proizvođača. Rezulati sekvenciranja su analizirani dostupnim softverom za analizu i prethodno razvijenim tokom rada (engl. pipeline). Interpretacija varijanti zasnovana je na analizi setova gena odabranih prema fenotipu bolesnika, pretraživanju literature i baza podataka, učestalosti alela, in silico analizama. Uzročne varijante su potvrđene sekvenciranjem po Sangeru i segregacionim analizama. Utvrđen je verovatan genetički uzrok kod devet nesrodnih bolesnika (četiri sa kognitivnim poremećajem i pet sa poremećajem pokreta). Detektovano je ukupno 11 patoloških varijanti u sedam gena (PSEN1, OPTN, TUBB4A, PANK2, SETX, MFSD8 i ARSA) od kojih su devet varijante sa promenjenim smislom (engl. missense), jedna varijanta bez smisla (engl. nonsense) i jedna varijanta u mestu splajsovanja (engl. splice site). Sve detektovane varijante su u genima koji su u skladu sa kliničkim fenotipom datih bolesnika. Pored toga, detektovane varijante u genima DCTN1, PDGFRB i POLG predstavljaju moguć uzrok bolesti kod tri dodatna slučaja sa poremećajem pokreta. Kod ostalih ispitanika genetička osnova bolesti ostaje nerazjašnjena. Rezultati ove teze ističu značaj analize panela klinički egzom u dijagnostici kognitivnih poremećaja i poremećaja pokreta i daju nam uvid u složenost genetičke pozadine ovih oboljenja.

Neurodegenerative diseases are characterized by a variable clinical picture, disease onset, prognosis and response to treatment. The largest number of them includes motor and cognitive disorders such as Alzheimer's and Parkinson's diseases, which are widespread, to extremely rare ones such as Creutzfeldt-Jakob disease. Here we analysed the genetic basis of cognitive and movement disorders by sequencing a clinical exome panel that comprise coding regions of 4813 genes with surrounding intronic sequencies in 57 unrelated patients (15 with cognitive and 42 with movement disorders). During selection, preference was given to family cases with an early onset of the disease or a complex phenotype. Panel sequencing was performed on the Illumina MiSeq platform according to the manufacturer's instructions. Results were analyzed with available software and an internal pipeline. The interpretation of variants is based on the analysis of gene sets selected according to the patient's phenotype, literature and database searches, allele frequencies, in silico analyses. Causal variants were confirmed by Sanger sequencing, as well as segregation analysis. A probable genetic cause was determined in nine unrelated patients (four with cognitive and five with movement disorder). We detected nine missense, one nonsense and one splice site pathological variants in seven genes (PSEN1, OPTN, TUBB4A, PANK2, SETX, MFSD8, and ARSA) which are in accordance with the clinical phenotype of the given patients. In addition, the detected variants in the DCTN1, PDGFRB, and POLG genes represent a possible cause of movement disorder in three additional cases. The rest of patients remain negative. The results of this thesis highlight the importance of clinical exome panel analysis in the diagnosis of cognitive and movement disorders and give us an insight into the genetic complexity of these diseases.