Targeted gene correction and functional recovery in achondroplasia patient-derived iPSCs

Achondroplasia (ACH) is the most common genetic form of dwarfism and belong to dominant monogenetic disorder (MGD) caused by a gain-of-function point mutation in the transmembrane region of fibroblast growth factor receptor 3 (FGFR3). There are currently two mutation sites reported – Gly380Arg and Gly375Cys, the former occupies the vast majority of ACH patients. Homozygous ACH patients have a much more severe phenotype and rarely survive, so most of the ACH patients seen are heterozygous mutations. Like many other MGDs, there are no effective therapeutic methods for ACH even though the cause of the mutation has been found. With the rapid development of stem cell biology and gene-editing technology, the research of ACH is not only important, but also provides theoretical and experimental basis for the investigation of other MGDs. In this study, we generated non-integrated induced pluripotent stem cell (iPSC) lines from ACH patient skin and urine of Gly380Arg mutation. Via powerful gene-editing tool – CRISPR-Cas9, we successfully obtained 2 completely corrected knock-in cell lines. They not only expressed pluripotent markers, maintained normal chromosomal number and structure, but also no off-target indels were identified. This is not only the first time to obtain ACH iPSCs using the most readily available biopsy sample – urine, the first to obtain non-integrated ACH iPSCs, but also is the first to perform gene correction in ACH research. Our study may provide important theoretical and experimental basis for the next step of stem cell research and treatment of ACH in the future.