A genetic tweak can make cells self-destruct in the presence of CRISPR and could be used to shut down cell therapies if they go rogue.
CRISPR gene editing can be used to efficiently and easily introduce changes to the DNA of living cells. It is a useful technique, but it would be handy to be able to make some cells CRISPR-resistant. For example, there is interest in storing information in DNA inside cells, and rendering some of these uneditable by CRISPR could enable us to make 鈥渞ead-only鈥� reference copies.
Cells that self-destruct in response to CRISPR could also be a useful brake for CRISPR-based gene drives 鈥� a technique that can rapidly spread detrimental mutations through a population, for example, to control or destroy a pest organism. Gene drives could be useful, but there are fears that the uncontrolled spread of genetic mutations could have harmful consequences. Making some organisms self-destruct when they encounter CRISPR could help control the spread of gene drives in the wild, says George Church at Harvard University.
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Genetic parasites
To make human cells CRISPR-proof, Church and his colleagues have exploited a genetic parasite that makes up around 17 per cent of our genome. Called LINE-1, this is a stretch of DNA that does nothing but make copies of itself. Our DNA contains tens of thousands of copies of it.
Standard CRISPR gene editing involves a protein called Cas9, which is given a guide RNA to find a specific DNA target sequence and then cuts the DNA at that site. Cells can repair a few DNA cuts, but if hundreds or thousands are made at the same time, their repair systems are overwhelmed and the cells self-destruct.
By engineering human cells in a dish to produce a guide RNA that targets LINE-1 DNA sequences, the team have made cells that die within days if anyone tries to edit them using the Cas9 protein.
Cell therapies
The team then took this a step further, to create a safety switch for cell therapies. Genetically engineered immune cells are starting to be used to treat several types of cancer, but they can trigger a deadly response called a cytokine storm. There are also worries that the immune cells themselves could turn cancerous.
But it should be possible to put a kill-switch into these cells. To do this, the team gave some human cells a gene to make the LINE-1 guide RNA, plus a gene for the Cas9 protein itself. However, this Cas9 gene had a regulatory sequence that meant that it could only become active in the presence of an antibiotic called doxycycline. When it was added to such cells in a dish, more than 99.9 per cent of them died within nine days.
The idea is that introducing this mechanism to immune cells before infusing them into a patient would give doctors a way to shut down any gene-edited cell therapy in the event of a cytokine storm or the cells turning cancerous. 鈥淚t could stop cell therapies that develop troublesome properties,鈥� says Church.
Fully tamper-proof
While there are already several other methods for creating cell safety switches, the researchers say theirs is better.
Kevin Esvelt at the Massachusetts Institute of Technology isn鈥檛 convinced the team鈥檚 switch would be much use for containing gene drives released accidentally or as bioweapons. The best way to counter a gene drive is to release another gene drive that reverses the first one, he says.
To ensure cells are fully tamper-proof, it would be necessary to protect against other kinds of proteins used in different flavours of CRISPR gene editing besides Cas9, says Carina Nieuwenweg at Wageningen University & Research in the Netherlands.
The team says this can easily be done with minor tweaks to the guide RNAs.
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