Excited about CRISPR? What if we’re all immune?

Image from flickr user ~db~ under a Creative Commons CC BY-NC-ND 2.0 Licence

In a paper published this week on the Biology preprint server bioRxiv (which publishes papers submitted by authors, which are not yet published in traditional journals), Researchers at Stanford have investigated human immunity to the key protein in the CRISPR genome editing system. This signal some challenges to the use of CRISPR as a therapy.Let’s back up a bit. CRISPR genome editing uses a bacterial system that exists as a kind of immune system to protect them from viruses. In has two components, a protein called Cas9 and a piece of the DNA-like polymer RNA called a guide RNA. The guide RNA is the targeting component of CRISPR, it binds to a specific DNA sequence, then the Cas9 cuts the DNA, often resulting in a mutation. We can use CRISPR simply to generate a mutation, or we can insert another piece of DNA.

From sequenced genomes, CRISPR systems are found in around 45% of bacteria and 85% of archae, but the two most commonly used in research are those from Staphylococcus aureus and Streptococcus pyogenes. These two species of bacteria are both commonly found on the skin and are often harmless, though can also be pathogenic, so it is likely that many people will have had some level of infection by these bacteria and have developed an immune response to many proteins from them. In this paper, the researchers investigated whether there is an immune response to the Cas9 proteins from these two species of bacteria.

To test for antibodies against Cas9, the researchers used serum from donated umbilical cords and then backed up their findings in serum from healthy adults. They found that many of the volunteers had antibodies against the two version of Cas9, with 86% of the umbilical cord samples, and 67% of the samples from adults having antibodies against the Staphylococcus aureus Cas9, and 73% of the umbilical cord samples and 42% of the adult samples having antibodies against the Streptococcus pyogenes Cas9.

The researchers then tested whether a specific immune cell called a T-cell (so called because they mostly mature in the thymus) from the blood of the adult volunteers also recognised the two versions of Cas9. They found some T-cells recognising the Streptococcus pyogenes Cas9, but the results were inconsistent. The results were clearer for the Cas9 from Staphylococcus aureus however, showing that 46% of donors had T-cells recognising this version of Cas9.

But what does this mean for CRISPR as a therapy? Well, CRISPR is being tested in two different ways as a therapy. So called ex vivo techniques extract cells from the patient, do the CRISPR genome editing, then reinject the cells. In a current small scale trial at a hospital in china, scientist are extracint immune cells from lung cancer patients blood, then using CRISPR to disable a protein called PD-1 which can restrict immune responses to cancer cells. The cells will then be injected back into the patient. In this kind of therapy, there won’t be a problem with an immune response to Cas9. In in vivo CRISPR, on the other hand, the Cas9 and guide RNA and targeted to the cells in the body. For example, in a trial for targeting the common virus HPV, which can cause cervical cancer, DNA coding for the Cas9 and guide RNA will be provided in a gel applied to the cervix. The plan is that the CRISPR will destroy the viral genes of infected cells, removing the increased cancer risk.

The presence of T cells against Cas9 is a concern for this kind of treatment. There are two potential problems to an immune response to a treatment such as this. The first is that the immune response will just stop the treatment from working. The second more worrying possibility is that if a larger immune response happens, there could be a systemic inflammatory response, making the patient ill.

So does this mean that we will have to right off in vivo CRISPR as a therapy? Not quite yet. I think this is more of a hurdle than an intractable problem. It might be possible to use Cas9 from another species of bacteria. Or we might be able to modify either the delivery method or the Cas9 itself to remove the immune response. CRISPR therapy is likely to be a big deal in the future and there are numerous clinical trials coming soon, so I think people are bound to work a way around this problem.


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