Patient-iPSC-Derived Kidney Organoids Show Functional Validation of a Ciliopathic Renal Phenotype and Reveal Underlying Pathogenetic Mechanisms

Despite the increasing diagnostic rate of genomic sequencing, the genetic basis of more than 50% of heritable kidney disease remains unresolved. Kidney organoidsdifferentiated from induced pluripotent stem cells(iPSCs) of individuals affected by inherited renal disease represent a potential, but unvalidated, platform for the functional validation of novel gene variants and investigation of underlying pathogenetic mechanisms. In this study, trio whole- exome sequencingof a prospectively identified nephronophthisis(NPHP) probandand her parents identified compound-heterozygous variants in IFT140 , a gene previously associated with NPHP-related ciliopathies. IFT140 plays a key role in retrograde intraflagellar transport, but the precise downstream cellular mechanisms responsible for disease presentation remain unknown. A one-step reprogramming and gene-editing protocol was used to derive both uncorrected probandiPSCs and isogenic gene-corrected iPSCs, which were differentiated to kidney organoids. Proband organoidtubules demonstrated shortened, club-shaped primary cilia, whereas gene correction rescued this phenotype. Differential expression analysis of epithelial cells isolated from organoidssuggested downregulation of genes associated with apicobasal polarity, cell- cell junctions, and dyneinmotor assembly in probandepithelial cells. Matrigelcyst cultures confirmed a polarization defect in probandversus gene-corrected renal epithelium. As such, this study represents a "proof of concept" for using proband-derived iPSCs to model renal disease and illustrates dysfunctional cellular pathways beyond the primary ciliumin the setting of IFT140 mutations, which are established for other NPHP genotypes.