Rational design of a Lfng-enhancer AAV construct drives specific and efficient gene expression in inner ear...

Area of Science:

• Oto-genetics
• Molecular Biology
• Gene Therapy

Background:

• Cell-specific gene expression is vital for effective gene therapies targeting sensorineural hearing loss.
• Current adeno-associated virus (AAV) serotypes have limited specificity for inner ear supporting cells.

Purpose of the Study:

• To develop a gene therapy tool for specifically targeting inner ear supporting cells.
• To combine an AAV serotype with high inner ear tropism and supporting cell-specific enhancer sequences.

Main Methods:

• Bioinformatic analysis identified three candidate Lunatic Fringe (Lfng) enhancer sequences.
• Adeno-associated virus (AAV)-ie capsids were engineered with Lfng enhancers or a CBh promoter driving EGFP.
• Vectors were introduced into neonatal mouse inner ears to assess gene expression and cell targeting.

Main Results:

• One Lfng enhancer demonstrated robust EGFP expression in cochlear and vestibular supporting cells without targeting brain cells.
• AAV-Lfng-CreERT2 vectors induced strong recombination in supporting cells of reporter mice post-tamoxifen treatment.
• The developed tool specifically targets supporting cells in juvenile and adult inner ears.

Conclusions:

• A novel AAV-based tool enables precise targeting of inner ear supporting cells.
• This advancement facilitates the development of safer and more effective gene therapies for hearing loss.
• The Lfng enhancer provides a valuable resource for inner ear research and therapeutic strategies.