Engineering and Selection of Shuffled AAV Genomes: A New Strategy for Producing Targeted Biological Nanoparticles

Area of Science:

• Molecular Biology
• Virology
• Gene Therapy

Background:

• Adeno-associated viruses (AAVs) are promising vectors for gene therapy but often lack cell type specificity.
• Developing targeted AAV vectors is crucial for efficient and safe gene delivery.

Purpose of the Study:

• To create a novel, cell type-specific AAV vector using DNA shuffling and directed evolution.
• To enhance AAV tropism for melanoma cells and evaluate its performance in various tissues.

Main Methods:

• DNA shuffling of AAV serotypes 1-9 capsid genomes to create a chimeric library.
• Isolation and characterization of a specific infectious clone (chimeric-1829).
• Molecular modeling, rational mutagenesis, and in vivo tropism studies.

Main Results:

• Chimeric-1829, derived from AAV1, 2, 8, and 9, efficiently transduces melanoma cells, utilizing heparan sulfate as a primary receptor.
• The C-terminal domain of AAV9 was identified as critical for melanoma tropism.
• Altered tropism was observed in rodent and nonhuman primate tissues (skeletal muscle, liver, brain).
• Unique immunological profile and crossreactivity were determined.

Conclusions:

• DNA shuffling is an effective method for generating targeted AAV vectors.
• Chimeric-1829 represents a promising synthetic capsid variant for gene transfer, with potential applications in human gene therapy.
• This approach can be extended to develop novel biological nanoparticles for diverse cell/tissue types.