Background <p>Nephronophthisis (NPH) is an autosomal recessive kidney disease, and <i>NPHP1</i> is the most frequently affected gene. Tubulointerstitial fibrosis is the major phenotype of <i>NPHP1</i>-deficient NPH. The pathophysiology of <i>NPHP1</i>-deficient NPH is unclear because models representing the disease pathophysiology are lacking. Herein, we aimed to create a novel pathological model of NPH using 3D kidney organoids derived from human-induced pluripotent stem cells (iPSCs) and elucidated the pathophysiology while searching for therapeutic candidates.</p> Methods <p><i>NPHP1</i>-deficient kidney organoids were generated from iPSCs. Fibrosis was induced by treatment with IL-1β. The effects of the Hippo signaling pathway inhibitors as therapeutic candidates were assessed. Fibrotic status was evaluated using immunofluorescence and quantitative PCR.</p> Results <p><i>NPHP1</i>^{<i>−/−</i>} kidney organoids were generated from iPSCs. Fibrosis induction with IL-1β considerably increased the expression of fibronectin and transcription of fibrosis-related genes in <i>NPHP1</i>^{<i>−/−</i>} organoids. Long-term culture of <i>NPHP1</i>^{<i>−/−</i>} organoids induced substantial fibrogenesis compared with wild-type organoids. Co-immunoprecipitation analysis revealed the binding of NPHP1 to LATS1/2—the main constituents of the Hippo pathway. IL-1β administration increased the expression of the key Hippo pathway genes in <i>NPHP1</i>^{<i>−/−</i>} organoids. By contrast, the Hippo pathway inhibitors ameliorated IL-1β-induced fibrogenesis in <i>NPHP1</i>^{<i>−/−</i>} organoids. Because one of the inhibitors, verteporfin, is in clinical use, its practical availability is expected from a drug-repositioning perspective.</p> Conclusions <p>Hippo signaling pathway is involved in the fibrotic changes associated with <i>NPHP1</i>-deficient NPH and the Hippo pathway inhibitors could be therapeutic agents.</p> Clinical trial number <p>Not applicable.</p>