Senior Director Thermo Fisher Scientific Carlsbad, California, United States
Abstract: Rare diseases, often genetic in origin, are complex and pose significant challenges in diagnosis and treatment. Zhu-Tokita-Takenouchi-Kim Syndrome (ZTTK) is one such rare condition caused by de novo heterozygous mutations in the SON gene, leading to haploinsufficiency and loss-of-function. It is characterized by intellectual disability and developmental delays. Developing treatments for rare diseases is resource-intensive and hindered by limited scientific understanding and inadequate models. Induced pluripotent stem cell (iPSC) technology offers substantial advantages as patient-specific, genetically accurate disease models for drug study and screening. In this study, we report the first successful generation of iPSCs from a 2-year-old ZTTK patient with a 4-bp (TTAG) deletion in the SON gene, causing a frameshift mutation and nonsense-mediated mRNA decay. Additionally, normal iPSCs were gene-edited to introduce the 4-bp mutation, creating syngeneic control lines. Peripheral blood from a ZTTK-affected infant and a healthy adult donor was reprogrammed using the CytoTune-iPS 2.0 Sendai Reprogramming Kit. iPSCs were successfully generated and banked. The 4-bp mutation was introduced into normal iPSCs via precise genome editing. Three genome-editing approaches—Cas9, Cas12a, and TALENs—were evaluated for optimal target recognition and knock-in efficiency. A streamlined process for clonal isolation and expansion of iPSCs and edited cells was established. Generated ZTTK-iPSCs and edited WT-iPSCs with the 4-bp deletion were characterized to confirm pluripotency, normal karyotype, and mutation presence. The iPSC clones produced in this study will be cryopreserved, deposited in a cell bank, and made available to researchers as valuable models for ZTTK syndrome research.
