Anomalies affecting the canine tooth include agenesis or impaction and the aetiology is still largely undetermined, with both genetic and environmental factors assumed to contribute significantly to the phenotypic manifestations. Recently, multiple lines of evidence highlighted the role of a few specifc cell signaling pathways, e.g. the NF-kB and the WNT, as crucial mediators of tooth development and tooth agenesis and inclusion. This evidence has been recently confirmed by the applicant through the clinical and genetic study of an Italian family with several members with maxillary canine anomalies, including canine agenesis, either monolateral or bilateral, canine impaction and canine ectopic eruption, characterized by incomplete penetrance and variable expressivity. The use of whole exome sequencing (WES) approach to characterized molecular bases of this heterogeneous condition highlighted the role of WNT and EDA cell signaling pathways in tooth development.
The aim of this research project is to study molecular bases of maxillary canine anomalies, including agenesis and impaction, in sporadic and familiar cases through a detailed clinical characterization and a Next GenerationSequencing approach.
The Department of Orthodontics at Sapienza University of Rome (Director: Professor Ersilia Barbato) is a reference center for patients with canine anomalies. The prevalence of canine impaction at the Orthodontics Unit is about 9%, with 75% of canine impaction anomalies affecting maxillary canines (Mercuri et al., 2013). This prevalence gives the opportunity to investigate the genetic component of canine anomalies on a large and well characterized sample.
A detailed clinical characterization of both sporadic and familiar cases has been already collected. Several cases has been already collected. Specifically, an interesting family with 7 family members with variable canine anomalies (impacted maxillary canine, either monolateral or bilateral, labial or palatal) has been already analysed and collected. Family history, clinical evaluation, radiographic and photographic examination were performed and blood samples were collected for 5 family members. The applicant plans to performed a WES approach on this and further familiar cases of canine anomalies.The occurrence of several subjects in a single family with anomalies affecting only maxillary canine without additional anomalies involving other teeth (e.g. agenesis of the third molars) is a very rare condition, therefore the study of this kind of pedigrees represent a valuable opportunity to investigate the molecular bases underlying this phenotype spectrum.
The results of this kind of study could support to solve a thorny issue that has been debated for about 20 years dealing with the different theories on the etiology of maxillary canine anomalies (Becker and Peck, 1995; Becker and Peck, 2016). This has been recently confirmed by the applicant through the characterization of the molecular bases underlying a complex trait consisting of isolated canine anomalies in a medium-sized family, highlighting the role of WNT and EDA cell signaling pathways in tooth development (Barbato et al. 2018).
Dissecting the genetic component underlying canine anomalies is a challenging task, as there are several limiting factors that influence gene identification, for example complex inheritance, reduced penetrance, variable expressivity, and the occurrence of sporadic cases that makes linkage analysis unfeasible.
Over the last few years, progresses in DNA sequencing technologies allowed to overcome these limitations, changing the way to study genetic diseases and genetic bases of complex traits leading to identification of molecular bases of human disease at an accelerating rate (Boycott et al., 2013). These technologies, known as "massive parallel sequencing" or "next generation sequencing" (NGS), provide information on a genomic scale, with a single base resolution, in a rapid and cost effective way (van Dijk et al., 2014). Whole
exome sequencing approach captures and sequences all the coding portion of the human genome (about 1%). Currently it is the most used approach for the discovery of rare-disease-causing genes as it has been estimated that 85% of the disease-causing mutations are located in the coding portion of the human genome, and because its feasibility in terms of computational infrastructures, costs, and
interpretation effort. WES is considered the most suited strategy for discovering rare alleles underlying Mendelian phenotypes and complex traits, as well as predisposing genetic variants in common diseases and cancers. Since tooth formation is a complex regulatory network with several genes potentially involved, we selected WES as the most effective strategy to dissect the genetic component related to canine anomaly spectrum. To date, only very few papers reported the use of NGS strategy to study genetic etiology of orodental anomalies, in particular canine anomalies (Prasad MK et al., 2016, Ockeloen CW et al., 2016, Massink MP et al., 2015, Salvi et al., 2016, Yamaguchi et al., 2017, Barbato et al. 2018).
Elucidating molecular bases causing tooth agenesis is a fundamental requirement for genetic counseling and also for the clinical management of dental anomalies, in order to prevent occlusion problems, particularly during developing dentition in children.