Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is a dominant genetic disorder caused primarily by mutations in the PKD1 gene, resulting in the formation of numerous cysts and eventually kidney failure. However, there are currently no gene therapy studies aimed at correcting PKD1 gene mutations. In this study, we identified two mutation sites associated with ADPKD, c.1198 (C > T) and c.8311 (G > A), which could potentially be corrected by adenine base editor (ABE). The correction efficiencies of different ABE variants were tested using the HEK293T-PKD1 c.1198 (C > T) and HEK293T-PKD1 c.8311 (G > A) reporter cell lines. We then generated induced pluripotent stem cells (iPSCsmut/WT) from the peripheral blood mononuclear cells (PBMCs) of the heterozygous patient to develop a disease cell model. Since the iPSCsmut/WT did not exhibit a typical disease phenotype in stem cell status, differentiation into kidney organoids in vitro led to the expression of kidney organ-specific marker proteins. Stimulation of cAMP signaling with forskolin resulted in cystic expansion of renal epithelial tissue in iPSCmut/WT-derived kidney organoids, resembling the cystic phenotype observed in ADPKD patients. However, kidney organoids differentiated from ABE-corrected iPSCs did not display the cystic phenotype. Furthermore, we used a dual AAV split-ABEmax system as a therapeutic strategy and achieved an average editing efficiency of approximately 6.56% in kidney organoids. Overall, this study provides a framework for gene therapy targeting ADPKD through ABE single-base editing, offering promising prospects for future therapeutic interventions.