A novel functional gene delivery platform based on a commensal human anellovirus demonstrates transduction in multiple tissue types

Anelloviridae is a family of non-enveloped viruses with negative-sense, circular, single-stranded deoxyribonucleic acid (ssDNA) genomes that infect vertebrates and are a ubiquitous component of the human virome. Human anelloviruses evade induction of humoral immune responses and appear to be non-pathogenic. These properties, in conjunction with their enormous genomic diversity and wide tissue distribution, make anelloviruses compelling candidates as vectors for next-generation genetic medicines. Here we report the first gene delivery vector system based on a human commensal virus. This Anellovector is based on a virus of the Betatorquevirus genus. Production is enabled by the development of the Self-Amplifying Trans-complementation of a Universal Recombinant aNellovector (SATURN) system, which relies on a self-replicating plasmid to provide viral proteins in trans that drive replication and capsid-dependent packaging of vector genomes. The SATURN system also utilizes a Cre-lox-based recombination mechanism to generate single unit-sized circular genomes inside the MOLT-4 production cell line. We demonstrate that the SATURN system can package a vector genome from a single betatorquevirus with capsids from multiple betatorquevirus species, supporting the feasibility of establishing a novel vector platform that takes advantage of the remarkable diversity of anelloviruses. The Anellovector demonstrated function in vitro in retinal pigment epithelial (RPE) cells. The Anellovector also demonstrated durable in vivo function in the mouse eye for 9 months after subretinal administration, and achieved comparable gene expression to dose-matched adeno-associated virus 9 (AAV9) when transduced by the intracerebroventricular (ICV) route of administration. To our knowledge, this is the first report of a functional anellovirus-based gene therapy vector. Anellovectors have great potential to deliver safe, redosable, and potent therapeutics, helping to expand the reach of programmable medicines.