Pre-arrayed Pan-AAV Peptide Display Libraries for Rapid Single-Round Screening.

Area of Science:

• Molecular Biology
• Gene Therapy
• Virology

Background:

• Displaying peptides on adeno-associated virus (AAV) surfaces is key for developing gene therapy vectors with improved cell targeting.
• While AAV serotype 2 (AAV2) is well-studied, other AAV variants show promise as scaffolds for peptide display.

Purpose of the Study:

• To systematically evaluate the potential of 13 different AAV capsid variants for displaying 27 peptides.
• To identify superior AAV vectors for gene therapy applications across diverse cell types.

Main Methods:

• Production of reporter-encoding AAV vectors with peptides inserted on capsid surfaces.
• Single-round screening of vectors across more than 90 cell types, including T cells and primary cells.
• Assembly of all experimental results into a searchable online database.

Main Results:

• Identified superior AAV vectors by screening peptide insertions across multiple AAV capsids and cell types.
• Demonstrated that vector performance is influenced by capsid type, peptide, cell type, insertion site, and flanking residues.
• Established a high-throughput reverse-transduction pipeline for efficient library screening.

Conclusions:

• Alternative AAV capsids offer vast potential for peptide display, accelerating the development of targeted gene therapy vectors.
• The findings provide a comprehensive resource for selecting and engineering AAV vectors for specific in vivo applications.
• This work facilitates the screening and application of novel AAV-based gene therapy tools.