Gene editing using gamma modified PNA: HBB gene as a model

PNAs have emerged as a powerful tool in gene editing, particularly for correcting monogenic disorders by enhancing targeted recombination and genomic modifications. This study aimed to establish a gene editing technique using Peptide Nucleic Acid (PNA)/donor DNA-loaded poly lactic-co-glycolic acid (PLGA) nanoparticles at our genomic facilities, with the ultimate goal of correcting disease-causing mutations. Methods involved culturing skin fibroblasts from a healthy Egyptian volunteer without HBB gene mutations in two separate 12-well plates. Oligonucleotides were designed, and nanoparticles were formulated and characterized before being used to treat the cultured fibroblasts. DNA and RNA were extracted from treated cells, followed by molecular analyses to confirm the edits. Results indicated successful encapsulation of nanoparticles and modest, sustained introduction of the desired mutation, accompanied by functional impairment in HBB gene expression. The study successfully established PNA gene editing technology, potentially paving the way for future treatment studies of single-gene disorders.