Primary ciliopathies are a group of rare inherited disorders caused by defects in the structure or function of primary cilia (the ‘cell’s antenna’). This thesis describes approaches to improve molecular diagnosis rates for primary ciliopathy patients over the ~40-80% currently achieved, through whole genome sequencing (WGS) analysis and functional variant interpretation.
Firstly, I analysed WGS data from the 100,000 Genomes Project (100K) for participants who were clinically suspected to have primary ciliopathies. I identified a molecular diagnosis rate for n=45/83 (54.2%), providing a 21.7% diagnostic uplift compared to results previously reported by Genomics England (GEL).
I then performed a reverse phenotyping study, starting by looking for pathogenic variants in nine multisystemic ciliopathy disease genes across the 100K rare disease dataset. This was linked back to available clinical data, aiming to identify participants with “hidden” ciliopathy diagnoses recruited to alternative categories. I identified 18 new, reportable diagnoses and 44 previously reported by GEL. I also found 11 un-reportable molecular diagnoses, lacking key clinical features to provide a confident fit for phenotype. This shows that the quality of entered phenotypic data is critical to allow accurate genotype-phenotype correlation.
In a third study, I developed strategies for functional interpretation of eight TMEM67 missense variants of uncertain significance (VUSs) with collaborators in Ireland, using CRISPR/Cas9 gene editing in a human ciliated cell-line (RPE-1) and C. elegans. These assays provided interpretation of three VUS as benign and five as pathogenic.
The two 100K studies show that diagnosis rates for ciliopathies can be improved through WGS analysis, especially structural and splice variant analysis. We are a long way from delivering a high-throughput system for VUS interpretation that could provide clinical utility in the diagnostic setting. Overall, we have provided benefit for ciliopathy patients through additional molecular diagnoses, accompanied by transferable skills applicable to wider patient groups
