Evaluating clinical significance of gene dosage sensitivity for kidney stone disease

Principle Investigator

Scientific interest within the context of the graduate college:

Our group is interested in the identification and investigation of genetic, clinical, and environmental factors determining the onset of chronic kidney disease (CKD) and kidney survival. We make use of next-generation sequencing techniques and deep-phenotyping to identify genetic variants that are predictive for disease progression or convey protection from organ failure. We functionally evaluate identified germline variants in vitro in order to understand underlying molecular mechanisms leading to CKD on the one hand or protecting from kidney failure on the other. By doing so, we aim at defining and targeting molecular switches responsible for health maintenance and disease alleviation.

Project description:

Introduction: Nephrolithiasis, or kidney stone disease (KSD), is a common global healthcare problem with a lifetime prevalence of 10-20%. Kidney stones recur frequently and cause substantial morbidity –including CKD, reduced quality of life and enormous cost. Kidney stone formation is strongly influenced by genetic factors (positive family history in 30-60%) and >30 Mendelian forms of KSD were described. Yet, molecular mechanisms of stone formation remain unknown in many adult kidney stone formers (KSF). Increasing data suggests relevant genetic risk for KSD from heterozygous variants in genes traditionally believed to have recessive inheritance like SLC34A3,^1,2 a phenomenon that is called gene dosage. We recently performed exome sequencing (ES) of a large cohort of 787 adult KSF and 114 non-KSF to assess prevalence of genetic disease in adult stone formers. Thereby, we detected a relatively low prevalence of Mendelian disease, but enrichment of genetic variants predisposing to KSD, amongst others, a heterozygous variant in CLDN16,³ encoding a renal tight junction protein. Homozygous pathogenic variants in CLDN16 (Claudin 16) cause a severe disease manifesting in childhood – often leading to kidney failure.⁴ Data suggest that heterozygous variants in CLDN16 may cause a less severe, but intermediate phenotype associated with an increased risk of KSD and CKD.⁵

Aim 1/WP1: Identification of genetic variants with potential gene dosage effect relevant for KSD. The applicant will perform detailed genotype-phenotype correlation of variants in CLDN16 inthe given dataset and compare results with a validation cohort of 800 KSD and with data in public repositories (e.g., Genomics England, UK Biobank, AllofUs). A similar approach assessing other rare and common variants (identified by single variant analysis and aggregated variant burden analysis) will be conducted for other candidate KSD genes with suspected gene dosage effect to identify genes and variants suitable for functional analysis (WP2).

Aim 2 /WP2: Functional validation of genetic variants with potential gene dosage effect in KSD. Initially, heterozygous CLDN16 variants found with increased prevalence (e.g., enriched) in stone formers will be studied. In a second step, a limited number of candidate genes and variants identified in WP1 will be assessed. For CLDN16, variants will be characterized in vitro in an established overexpression system in collaboration with the laboratories of Prof. Dorothee Günzel and PD Dr. Jörg Piontek, both known experts in tight junction proteins. Additionally, depending on the results from overexpression studies, characterization of urinary renal epithelial cells isolated from selected individuals with the respective CDLN16 variants will be performed. Planned analyses include qRT-PCR, Western Blot, immunofluorescence imaging and measurement of transepithelial resistance (TER).

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