Nat Biotech: Producing 1 billion CAR-T cells at a time, the new CRISPR technology efficiently inserts ultra-long DNA sequences without the need for viral vectors

The advent of CRISPR technology has significantly changed the landscape of gene editing, providing scientists with unprecedented ease and efficiency

Conventional CRISPR technology usually relies on viral vectors to deliver to cells, but viral vectors are often expensive and resource-intensive, so manufacturing a large number of clinical-grade viral vectors has always been one

Recently, researchers at the University of California, San Francisco, published a research paper

The study developed an improved CRISPR-Cas9 gene editing technology that can very efficiently introduce long DNA sequences into the precise location of the cell genome without the need for a viral vector, achieving 2-3 times higher editing efficiency

In classical CRISPR-Cas9 gene editing techniques, high concentrations of double-stranded DNA (dsDNA) and Cas9 target sequences are required to enhance CRISPR-mediated insertion efficiency, but this can be more toxic

In addition, to overcome the drawbacks of the need for viral vectors by conventional CRISPR techniques, for years the research team has been working to insert longer DNA sequences into specific sites in the genome in a way that does not rely on viral vectors, and to enable more efficient gene editing

In this latest study, the research team developed a new CRISPR system that utilizes a single-stranded DNA (ssDNA) HDR template (HDRT) that binds to the Cas9 target sequence to achieve 2-3 times the knock-in efficiency and yield

Development of a new CRISPR system

Not only that, but the research team also showed how the new CRISPR system can be used to quickly produce CAR-T cells

Using a new DNA template, the research team generated more than 1 billion CAR-T cells that target multiple myeloma, and about half of them acquired new genes and converted to CAR-T cells

Utilize the new CRISPR system to rapidly produce CAR-T cells

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The new CRISPR system can design more than 1 billion CAR-T cells in a single run

In addition, the researchers showed for the first time that their approach could completely replace two genes associated with rare inherited immune diseases, namely IL2RA and CTLA4

Full ORF replacement of therapeutic and diagnostic target genes with human T cell editing

What's more, the findings show that nearly 90 percent of the cells treated with this genetic engineering method acquired healthy versions of

Overall, the study, published in Nature Biotechnology, develops a novel, vector-independent CRISPR-Cas9 gene-editing system that achieves 2-3 times higher editing efficiency

Original source:

Shy， B.