Varsha Jonnalagadda

Assessing CRISPR Guide Site Viability and Downstream Transcriptomic Effects in CFTR Editing

Started June 2, 2025

Abstract or project description

Cystic fibrosis (CF) is a life-threatening genetic disorder caused by mutations in the CFTR gene, which encodes a chloride channel essential for fluid regulation in the lungs and other organs. With CRISPR/Cas9 gene editing positioned as a potential cure, this study critically evaluates both the technical feasibility of editing CFTR and the broader biological consequences of doing so. First, guide RNA (gRNA) target sites across all 26 exons of CFTR were analyzed using CHOPCHOP. The results reveal a striking lack of high-fidelity target sites, indicating that full-gene editing is not only difficult but likely unsafe with current tools. In the second phase, RNA sequencing data from human epithelial cells with a CFTR knockout were analyzed to uncover downstream effects. The transcriptomic changes were profound, disrupting key cellular pathways involved in immune regulation, lipid metabolism, detoxification, and the management of oxidative stress. One pathway - LPS/IL-1-mediated inhibition of RXR function - stood out for its widespread physiological consequences, linking CFTR loss to chronic inflammation, metabolic instability, and increased disease risk. These findings strongly suggest that CFTR is not only structurally resistant to precise gene editing but also plays a central role in maintaining biological stability. As a result, CRISPR-based editing of CFTR may carry substantial, system-wide risks, raising urgent questions about the safety and ethics of pursuing such interventions without a deeper understanding of their ripple effects. This research provides a critical framework for reassessing gene-editing strategies in the treatment of complex monogenic diseases.