Allele-Specific Knockdown of Mutant Huntingtin Protein via Editing at Coding Region Single Nucleotide Polymorphism...

Area of Science:

• Neuroscience
• Genetics
• Biotechnology

Background:

• Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder.
• It results from a trinucleotide repeat expansion in the huntingtin (HTT) gene.
• CRISPR-Cas9 gene editing is a potential therapeutic strategy, but permanent HTT disruption is a concern.

Purpose of the Study:

• To investigate allele-specific gene editing of mutant huntingtin (mHTT).
• To assess the feasibility of reducing mHTT protein without affecting normal HTT function.
• To evaluate CRISPR-Cas9 efficacy in a novel HD mouse model.

Main Methods:

• Utilized a novel HD mouse model with BAC97 (mHTT) and YAC18 (normal HTT) transgenes.
• Employed CRISPR-Cas9 gene editing targeting a single nucleotide polymorphism (SNP) for allele-specific inactivation.
• Analyzed mutation frequency (InDels, viral insertions) in the targeted HD allele within neurons.

Main Results:

• Achieved allele-selective reduction of mutant huntingtin protein in the HD mouse model.
• Demonstrated high-frequency mutations (InDels, insertions) in the targeted HD allele.
• Confirmed successful inactivation of the mutant HTT allele.

Conclusions:

• Allele-specific CRISPR-Cas9 editing targeting heterozygous SNPs is a feasible approach for inactivating autosomal dominant mutations.
• This strategy offers a potential therapeutic avenue for Huntington's disease by selectively targeting the mutant allele.
• Reduces mHTT protein levels while preserving normal HTT function, mitigating risks of permanent disruption.