Rescue of premature aging defects in Cockayne syndrome stem cells by CRISPR/Cas9-mediated gene correction

Cockayne syndrome(CS)is a rare autosomal recessive inherited disorder characterized by a variety of clinical features,including increased sensitivity to sunlight,progressive neurological abnormalities,and the appearance of premature aging.However,the pathogenesis of CS remains unclear due to the limitations of current disease models.Here,we generate integration-free induced pluripotent stem cells(iPSCs)from fibroblasts from a CS patient bearing mutations in CSB/ERCC6 gene and further derive isogenic genecorrected CS-iPSCs(GC-iPSCs)using the CRISPR/Cas9 system.CS-associated phenotypic defects are recapitulated in CS-iPSC-derived mesenchymal stem cells(MSCs)and neural stem cells(NSCs),both of which display increased susceptibility to DNA damage stress.Premature aging defects in CS-MSCs are rescued by the targeted correction of mutant ERCC6.We next map the transcriptomic landscapes in CS-iPSCs and GC-iPSCs and their somatic stem cell derivatives(MSCs and NSCs)in the absence or presence of ultraviolet(UV)and replicative stresses,revealing that defects in DNA repair account for CS pathologies.Moreover,we generate autologous GC-MSCs free of pathogenic mutation under a cGMP(Current Good Manufacturing Practice)-compliant condition,which hold potential for use as improved biomaterials for future stem cell replacement therapy for CS.Collectively,our models demonstrate novel disease features and molecular mechanisms and lay a foundation for the development of novel therapeutic strategies to treat CS.