Taming T4: Artificial Viral Vectors Deliver Big Payloads into Human Cells
In a new study published in Nature Communications, Rao and his team show how to construct artificial viral vectors (AVVs) using an assembly-line technique that allows for the delivery of large biomolecular payloads, such as 171 kilobases of DNA, thousands of protein molecules, and RNAs. In order to edit the human genome—recombine or replace genes, alter or silence gene expression—they have redesigned the well-characterized structural components of bacteriophage T4 to accomplish these tasks.
In order to program and deliver therapeutic biomolecules, Rao and colleagues used T4 bacteriophages to build AVVs with a large internal volume and a large external surface. Proof-of-concept experiments showed the viability of these AVVs for use in genome engineering by loading them with protein and nucleic acid cargo. The full-length dystrophin gene and other molecular operations to modify the human genome were successfully carried out in the lab using the platform. The AVVs have a high yield and low cost of production, and the nanomaterials have been shown to be stable for several months.
Rao hopes that this technique could eventually be used to treat a variety of common and rare disorders in humans, although more study is needed to determine its safety.
“Neither NSF nor NIH were really interested in funding phage research that enthusiastically,” said Rao. “But I always believed in phages because these are good model systems to really understand the basic mechanisms of DNA packaging, there’s a lot we can manipulate genetically, and they are inexpensive. I can do it with students and a small research lab with Petri dishes and LB media.”
In his team’s latest work, Rao demonstrates how they have optimized this technology in the Nature Communications report, demonstrating their ability to package massive amounts of DNA, RNA, and proteins, as well as complexes combining these biomolecules, such as CRISPR systems with guide RNAs.
While Rao’s team has demonstrated the feasibility of their system in immortalized cell lines like 293 T cells, they are currently working to implement it in primary human cells and human embryonic stem cells, and even in an in vivo animal model. The goal, according to Rao, is to get the system into a mouse model as soon as possible so that it can be brought to the clinic in the not-too-distant future.
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