🧫 Experiment Protocol
Clinicalproposed
SUMMARY
# CRISPR Gene Correction Approaches for CBS/PSP
## Background and Rationale
CRISPR-Cas9 gene editing represents a transformative therapeutic modality for treating monogenic forms of neurodegeneration, offering unprecedented precision for correcting disease-causing mutations at their genomic source. Progressive supranuclear palsy (PSP) and corticobasal syndrome (CBS) include several genetic variants caused by mutations in MAPT (microtubule-associated protein tau), GRN (granulin), and C9orf72 gene
METHODOLOGY NOTES
**Phase 1: Patient Recruitment and Screening (Months 1-6)**
• Recruit 60 patients with genetically confirmed CBS/PSP (MAPT, GRN, or C9orf72 mutations)
• Obtain informed consent for research participation and genetic analysis
• Collect detailed clinical assessments using PSP Rating Scale and CBS severity measures
• Extract peripheral blood mononuclear cells (PBMCs) and establish patient-specific iPSC lines
• Perform whole genome sequencing to confirm pathogenic variants
**Phase 2: CRISPR Design and Validation (Months 3-9)**
• Design patient-specific guide RNAs targeting pathogenic mutations using CHOPCHOP and Benchling
• Synthesize Cas9-RNP complexes with homology-directed repair (HDR) templates
• Test editing efficiency in patient iPSCs using digital droplet PCR and Sanger sequencing
• Validate off-target effects using GUIDE-seq and targeted amplicon sequencing
• Optimize electroporation conditions for >70% editing efficiency
**Phase 3: Neuronal Differentiation and Correction (Months