Importance of whole exome sequencing in diagnosis of diseases (the Libyan experience)

Abstract

The exome is the part of the genome formed by exons (Figure 1); the sequences which when transcribed remain within the mature RNA after introns are removed by RNA splicing. The exon covers between 1 and 2% of the genome, depending on species [1]. Whole-exome sequencing (WES) is a revolutionary technique, which targets the protein-coding regions of the genome and has proven success in identifying new causal mutations and major congenital anomalies for diseases of previously unknown aetiology. With a successful diagnostic rate approaching 25% for rare disease, its clinical utility is becoming increasingly popular [3]. WES consists in the capture, sequencing and analysis of all exons of all protein-coding genes in the human genome. Instead of analysing the whole genome, composed of roughly three billion base-pairs, WES focuses only on approximately 30 million base-pairs which are translated into functional proteins. It is believed that the mutations of genes responsible for the formation of the functional proteins are the most likely to have a severe direct phenotypic consequence [4]. Therefore, WES was suggested to be much less costly and more efficient method of identifying all possible mutations in genes, compared to other methods such as genome-wide association studies or whole-genome sequencing (WGS) [4]. Before the discovery of WES, many patients with genetic diseases were not given a specific diagnosis. The lack of a diagnosis can have considerable adverse effects for patients and their families, including failure to identify potential treatments, failure to recognize the risk of recurrence in subsequent pregnancies, and failure to provide anticipatory guidance and prognosis [5]. Therefore, WES is extensively used nowadays to diagnose novel diseases and find novel causative mutations for known disease phenotypes. We in our paediatric neurology unit in Tripoli Children Hospital, serving the west and south area of the country with a population of 1.5 million children below 15 years of age, are faced almost on a daily basis with patients of developmental delay difficult to diagnose in spite of doing the basic neuro-metabolic screen and imaging. Since 2010, our unite is cooperating with the research center of Dr. Joseph Gleeson laboratory for paediatric brain diseases, Howard Hughes Medical Institute, University of California San Diego USA. We were able to send blood from 100 families whom they have at least two affected children without a clear diagnosis, and we were able to find out the causative gene in about 60% and we discovered about 10 novel genes, four of them have been published in highly ranked international journals [6,7,8]. When a researcher is faced with a novel gene variant, he/she needs to prove that this gene mutation is pathological. This prove requires a lengthy testing procedures by doing functional testing on knocked-down mouse or zebra fish for the specific mutant gene. Unfortunately, this kind of research is lacking in Libya and we should put all needed efforts to establish these techniques in our research laboratories. Moreover, the researchers in Dr Gleeson Laboratory now started to study WES negative patients by doing WGS, which mean to study the rest of 99% of the genome that don’t code for protein and performing sequencing of RNA from cell line skin fibroblast from these families. By comparing the sequencing and levels of RNA from patient’s cells we can solve more cases of patients with genetic defects.