When the signal is lost: Improving molecular diagnostics of renal ciliopathies

Nephronophthisis is the most important monogenic cause of pediatric end-stage renal disease. The average diagnostic delay in children with nephronophthisis is 3.5 years due to aspecific presenting symptoms. In addition, genetic testing can confirm the clinical diagnosis in less than 50% of patients, which hampers prognostics and genetic counseling. The research described in this thesis is aimed at improving diagnostics, prognostics and clinical care for patients with nephronophthisis-related ciliopathies (NPH-RC). To improve early detection and prognostics, the Nephronophthisis Registry was created to identify presenting signs and symptoms and to investigate gene-phenotype associations in a cohort of 40 Dutch patients with NPH-RC. Persistent fatigue, polyuria and hypertension were identified as initial signs and symptoms of nephronophthisis. In addition, increased echogenicity was a more prevalent finding on renal ultrasound than cysts. Importantly, we showed that symptom onset can occur well into adulthood, which supports continuation of presymptomatic monitoring of kidney function in syndromic ciliopathy patients until at least age 30 years. Novel genetic variants and candidate genes for NPH-RC were identified. We described NEK1 as the first gene for oral-facial-digital syndrome type II and added support for nonsense-associated alternative splicing as a mechanism behind phenotypic heterogeneity in ciliopathies. Variants in NEK1 and the candidate ciliopathy gene KIF14 were functionally characterized in cell models. KIF14-deficient fibroblasts showed an increased number of mitotic cells and absence of KIF14 at the midbody, which could suggest a role of cytokinesis defects in the pathophysiology of KIF14-associated Meckel-Gruber syndrome. Finally, liquid chromatography-mass spectrometry (LC-MS/MS)-based proteomics analysis of urinary extracellular vesicles was used as a novel method for non-invasive diagnostic and prognostic biomarker discovery in patients with NPH-RC. We showed for the first time that proteomic profiles differentiate NPH-RC patients from healthy controls. Expression levels of discriminating proteins were associated with stage of chronic kidney disease, suggesting potential prognostic applications. When validated, these results will facilitate early detection and timely treatment of NPH-RC. Ultimately, the combination of deep phenotyping and different molecular techniques including next-generation sequencing, RNA-sequencing and proteomics will yield the best diagnostic, prognostic and overall clinical care for NPH-RC patients.