Document Type : Original Article(s)
Authors
1 Department of Stem Cells Technology and Tissue Regeneration, Faculty of Interdisciplinary Science and Technologies, Tarbiat Modares University, Tehran, Iran
2 Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
3 Pediatric Cell and Gene Therapy Research Center, Gene, Cell, and Tissue Research Institute, Tehran University of Medical Sciences, Tehran, Iran
Abstract
Background: Leukocyte adhesion deficiency (LAD) is a genetic immunodeficiency that severely impairs the leukocyte adhesion cascade and migration to the sites of infection and inflammation. As a result, the main consequence is recurrent and severe infections with bacteria and fungi, which can lead to death. The most common variant, LAD-I, is due to mutations in the integrin beta 2 (ITGB2) gene responsible for the β2 integrin (CD18) subunit. Corrective strategies based on gene therapy and genome editing are the most promising methods for treating genetic defects.
Methods: In the present experimental in vitro study conducted at Tarbiat Modares University, Tehran, Iran, in 2024, we designed a single-guide RNA (sgRNA) for the CRISPR/Cas9 system to target exon 6 of ITGB2 and performed validation experiments. We also investigated peptide nanocarriers as a potential strategy for delivering the CRISPR components. The PX458 plasmid encoding Cas9 and the designed sgRNA was delivered into HEK293T cells, followed by genome-editing analysis.
Results: Genome-editing analysis confirmed a measurable level of indel formation at the target locus. Furthermore, the results of MPG nanopeptide production and purification demonstrate the potential of peptide nanocarriers for genome-editing systems.
Conclusion: Overall, this work establishes a foundation for subsequent knock-in and gene-correction studies in clinically relevant immune cell models, paving the way for potential LAD-I gene therapy applications.
Highlights
Sevil Raji (Google Scholar)
Hossein Soltaninejad (Google Scholar)
Keywords
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