Generation of an in vitro model for Hunter syndrome using CRISPR/Cas9 technology

VAENA E; ORMAZABAL, MAXIMILIANO; PAVAN, ELEONORA; MUCCI, JUAN M.; CRIVARO, ANDREA; BONDAR, CONSTANZA; CECI, ROMINA; ROZENFELD, PAULA; DARDIS, ANDREA

Simposio; 19th WORLD Symposium Lysosomal Disease Network; 2023

Hunter syndrome (mucopolysaccharidosis type II) is a lysosomal disease, caused by the deficiency in the lysosomal enzyme iduronate 2-sulphatase (IDS). The disorder compromises different organs and tissues, including liver, spleen, bones, joints, heart, lungs and central nervous system (CNS). To date, available treatments only show effects at visceral level, with limiting effects in CNS, joints, bones and lungs. Although studies on in vitro and in vivo models of the disease provided some information about the molecular basis of the disorder, they do not fully explain the mechanisms involved in the disease pathogenesis in particular in tissues that are not easily accessible. Therefore, our group developed an in vitro cellular models of MPS II joint tissues by editing the SW982 cell line (human synovial fibroblast) using CRISPR/Cas9 technology, and two RNA guides targeting different regions of the IDS gene. After editing we performed single cell sorting and selected clones with deficient IDS enzymatic activity, two of them were further characterized showing the presence of pathogenic variants in the IDS gene.