Genome editing strategies to generate working models of polycystic kidney disease

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is predominantly caused by pathogenic variants in PKD1 with truncating variants generally associated with more rapid cyst formation. However, it is difficult to predict disease progression, particularly in the case of missense variants. In this context, our aim was to provide a proof of principle that it is possible to generate cellular models carrying specific PKD1 variants. We generated homozygous (Ex15^-/-) and heterozygous mutants (Ex15^+/-) starting from HEK293T cells. Then, the variant c.11614G > A p.(E3872K) was introduced into PKD1 WT and heterozygous clones, obtaining compound heterozygous (Ex15^+/-/Ex42^M/M) cell lines. Functional assays confirmed the pathogenicity of the c.11614G > A variant. Ex15^+/-/Ex42^M/M cells behaved similarly to Ex15^-/- with defective actin polymerization, diminished capacity to complete autophagic flux and reduced mitochondrial respiration. Interestingly, they showed a different behavior in terms of cell-cycle regulation, implying mutation-specific effects. This work provides a starting point for the generation of different ADPKD-related variants using base editors, with the goal of complementing the already available knowledge about genotype-phenotype correlations and potentially reclassifying C3 variants in pathogenic or non-pathogenic.