EVALUATION OF CRISPR/CAS9 GENOME-EDITING SYSTEM IN HUMAN STEM CELLS HSCS: THERAPEUTICS AND DIAGNOSTICS PROSPECTS

Abstract

Background: CRISPR/Cas9 genome-editing technology has revolutionized human stem cell (HSC) research, offering novel therapeutic and diagnostic applications. HSCs play a crucial role in regenerative medicine and genetic therapies due to their ability to self-renew and differentiate into various blood cell lineages. The precise genome-editing capability of CRISPR/Cas9 allows for targeted gene modifications, enabling the correction of inherited disorders, disease modeling and the discovery of novel biomarkers. Despite significant advancements, challenges such as off-target effects, delivery efficiency, and ethical concerns persist, requiring further research and optimization.

Objective: This systematic review evaluates the therapeutic and diagnostic potential of CRISPR/Cas9 genome editing in HSCs, focusing on its efficacy in gene correction for hematologic disorders, disease modeling and biomarker discovery.

Methods: A systematic review was conducted following PRISMA guidelines, analyzing studies published between 2015 and 2024. Literature searches were performed in PubMed, Web of Science, and Scopus using MeSH-aligned keywords. The inclusion criteria encompassed peer-reviewed studies utilizing CRISPR/Cas9 for gene modification in HSCs for therapeutic and diagnostic applications. Exclusion criteria included studies that lacked experimental validation, did not focus on HSCs, or were non-English publications. Out of 85 initially retrieved studies, 40 met the inclusion criteria, and 15 were selected for final synthesis.

Results: CRISPR/Cas9 gene-editing strategies in HSCs were categorized as gene knockout (53%), gene activation (40%), and dual knockout/activation (7%). Hematological disorders, including sickle cell anemia and beta-thalassemia, accounted for 35% of studies, demonstrating up to 90% correction in β-globin mutations. Neurodegenerative diseases constituted 20% of studies, where knockout of amyloid precursor protein (APP) in Alzheimer’s models resulted in a 60% reduction in plaque accumulation. Muscular dystrophy studies (10%) showed 75% improvement in dystrophin expression through gene activation. High-throughput CRISPR screening was employed in 15% of studies for biomarker identification. Despite promising outcomes, off-target mutations were observed in 28% of studies, and viral vector-based delivery methods were used in 65%, raising safety concerns.

Conclusion: CRISPR/Cas9 genome editing in HSCs presents a ground-breaking approach for treating genetic disorders and enhancing precision medicine. Its potential to correct disease-causing mutations, model complex disorders, and identify novel therapeutic targets is substantial. However, challenges in delivery methods, long-term safety, and ethical considerations remain barriers to clinical translation. Future research should focus on optimizing high-fidelity Cas9 variants, improving non-viral delivery methods, and addressing ethical concerns to ensure the safe and effective application of CRISPR/Cas9 in regenerative medicine.