Researchers have developed a highly efficient new gene-editing method that uses virus-based protein fragments. The method could be used to level up existing cell and gene therapies used to treat cancer and other diseases.
Using CRISPR technology to simply and efficiently modify genes has revolutionized biomedical research and treatment and provided a better understanding of the genetic basis of diseases.
CRISPR is a component of bacterial immune systems that can cut DNA; it’s been repurposed as a gene-editing tool. Scientists design a guide RNA to match the gene they want to edit and attach it to a CRISPR-associated protein (Cas). The guide RNA directs Cas to the target gene, where it acts like ‘molecular scissors,’ snipping the troublesome DNA.
But for all its benefits, the technology has difficulty entering primary cells, which are cells taken directly from living tissue or organs and grown in a lab. T cells, part of the body’s immune system, are examples of primary cells.
After finding that some viruses use protein fragments – peptides – to enter cells, researchers from the University of Pennsylvania tested whether they could use this method to introduce CRISPR gene-editing technology more effectively into primary cells.
“Current methods of getting CRISPR-Cas systems into cells, which include the use of carrier viruses and electric pulses, are inefficient for cells taken directly from patients (called primary cells),” said Shelley Berger, co-corresponding author of the study. “These methods also typically kill many of the cells they are used on, and can even cause broad unwanted changes in gene activity.”
The researchers used peptides to guide CRISPR-Cas9 and Cas12a molecules through the outer membrane of primary human and mouse cells and into their nuclei, where most of the cell’s DNA is located. They found that using a combination of two modified peptides, one found in HIV and one in influenza viruses, mixed with CRISPR-Cas molecules, had a gene-editing efficiency of close to 100%, was non-toxic and did not cause changes to gene expression.
Calling their novel approach peptide-assisted genome editing, PAGE, the researchers say it could be particularly useful in T cell-related therapies, such as chimeric antigen receptor (CAR) T cell therapy, which uses specially modified immune cells taken from a patient to treat blood cancer.
“This new approach has the potential to be a major enabling technology for engineering cellular therapies,” said E. John Wherry, another corresponding author of the study.
In addition to its use in cell and gene therapies, PAGE has broad applications say the researchers.
“The simplicity and power of the peptide-assisted concept suggests that it could potentially be adapted in the future for the delivery into primary cells of other genome-editing proteins, or even protein-based drugs,” said Junwei Shi, co-corresponding author of the study.
The study was published in the journal Nature Biotechnology.
Source: University of Pennsylvania
