An E. coli-based platform for the production and assembly of anellovirus vectors

Abstract

Gene therapy offers immense potential for treating various diseases, including cancer, immunodeficiencies, and cardiovascular conditions. The efficacy of gene therapy (GT) largely depends on the vector used for gene delivery. Viral vectors, while effective, pose risks including insertional mutagenesis, immune responses, and high manufacturing costs. Non-viral vectors, although safer and easier to produce, often exhibit lower transfection efficiency and weaker transgene expression. This highlights the need for novel, more efficient vectors. Among emerging strategies, bacteriophages are gaining attention as promising GT delivery vehicles due to their adaptability and safety profile. Filamentous phages like M13 have demonstrated potential as targeted gene delivery vectors. This study proposes constructing a single-stranded DNA (ssDNA) phage-based vector incorporating a eukaryotic gene cassette. By leveraging Ff phage replication mechanisms in E. coli, the study explores encapsidating ssDNA within anellovirus capsids. These small, ssDNA viruses, known for their ability to transfect diverse tissues, offer a safer alternative to conventional viral vectors. Through successful expression and assembly of anellovirus capsid proteins, ssDNA viral particles were produced ex vivo. This innovative E. coli-based anellovirus-phagemid system provides a promising, cost-effective platform for developing next-generation viral vectors in gene therapy.