Search

Summary Human induced pluripotent stem cells (hiPSCs) allow in vitro study of genetic diseases and hold potential for personalized stem cell therapy. Gene editing, precisely modifying specifically targeted loci, represents a valuable tool for different hiPSC applications. This is especially useful in monogenic diseases to dissect the function of unknown mutations or to create genetically corrected, patient-derived hiPSCs. Here we describe a highly efficient method for simultaneous base editing and reprogramming of fibroblasts employing a CRISPR-Cas9 adenine base editor. As a proof of concept, we apply this approach to generate gene-edited hiPSCs from skin biopsies of four patients carrying a Finnish-founder pathogenic point mutation in either NOTCH3 or LDLR genes. We also show LDLR activity restoration after the gene correction. Overall, this method yields tens of gene-edited hiPSC monoclonal lines with unprecedented efficiency and robustness while considerably reducing the cell culture time and thus the risk for in vitro alterations., Graphical abstract, Highlights • Simultaneous base editing and reprogramming efficiently generates corrected hiPSCs • RNA-delivered ABE edits the targeted loci without DSBs or detectable off-targets • The base-edited hiPSCs exhibit normal karyotype and pluripotency • Differentiated patient hiPSCs show phenotype restoration upon gene correction, In this study, Jalil and colleagues develop and test a method for simultaneous hiPSC reprogramming and RNA-delivered adenine base editing. Starting from primary patient-derived fibroblasts, this approach yields several on-target edited isogenic hiPSC lines, with remarkable speed and editing efficiency and without compromising their genetic integrity or their pluripotency quality. The gene editing corrects the cellular disease phenotype.