American Scientist

In 2012, Emmanuelle Charpentier codiscovered the CRISPR-Cas9 system, a genetic tool that is changing the fields of genomics, genetics, and genomic engineering. CRISPR-Cas systems consist of RNAs and proteins that recognize DNA of invading viruses and cut it up. (The name stands for Clustered Regularly Interspaced Short Palindromic Repeats, describing its genetic pattern.) For many bacteria and archaea, this system functions as an effective immune system, but in the lab it can be used for remarkably targeted genomic editing. This tool could have vast uses in medicine and industry, and could facilitate human genetic engineering. Charpentier spoke with associate editor Katie L. Burke about the current direction and future applications of her research.

When you began working on CRISPR, it was a little-known region in the genomes of some bacteria. What drew you to the CRISPR system, and when did you realize its importance?

When my lab started to work on CRISPR, we were interested in how bacteria, such as Streptococcus pyogenes, cause diseases. We were trying to decipher regulatory mechanisms that explain how the bacteria are virulent and how they survive in the human host. We looked at the genome sequence of S. pyogenes to see whether there were portions of it that produced regulatory RNAs. RNA is probably best known as a molecule involved in the translation of genes into proteins, but it can also regulate the expression of genes rather than coding for proteins. We found a number of RNAs, including those of the CRISPR system and one that we called tracer RNA that was located next to the CRISPR locus.

In 2006 and 2007, it was not clear what the CRISPR system was. We showed that it is a defense system that protects the bacteria from viruses, which are composed of DNA. There are different defense systems that bacteria have evolved to destroy these invading genomes. CRISPR is such a system, one that is a little bit similar to our adaptive immune system. The CRISPR system first recognizes the invading genome and once it does so, it memorizes it. Upon a second infection, it can destroy the foreign DNA.

When the invading genome enters the bacteria, there will be recognition of it via an RNA molecule that contains the signature of the phage. One of the CRISPR-associated proteins [Cas proteins] will cut the genome of the phage, which will be the dead end for its genome expression and multiplication.

There are different types of CRISPR systems, and CRISPR-Cas9 is the simplest. It’s composed of two RNAs [CRISPR] and one protein [Cas9]. Together, they cleave the invading DNA. This process is what we described in two papers, one published in Nature in 2011 and another published in Science in 2012.

What makes the CRISPR-Cas9 system especially attractive for genomic engineering and genome editing?

Even though we can sequence and decipher the genetic code of any organism, scientists are still very limited with regard to the tools for manipulating the genome specifically and precisely. There are some recombination methods that have been developed over the last 40 years, but they were not very precise. What we call genome engineering, and especially genome editing, came out a little more than 10 years ago, when there were some classes of enzymes that were discovered, the meganucleases.

Those nucleases were very attractive to scientists, because they could then precisely introduce a mutation on the genome of eukaryotic cells, or correct a mutation, change a gene, exchange a gene, or delete a sequence. However, each time the scientist wants to manipulate a specific sequence, he or she would have to engineer a new protein. This process is costly, takes time, does not always work, and is not efficient.

CRISPR-Cas9 is different. The component that recognizes the DNA to be manipulated is contained in a small RNA molecule. So, the biologist just has to manipulate this RNA, according to the target DNA sequence. This feature makes this tool cheap and fast to design. It is efficient and has been shown to work in any cell and organism. In contrast to the meganucleases, which could only introduce mutations or exchange genes, CRISPR-Cas9 can also modify the DNA and be manipulated as a tool that will change the expression of the DNA. It’s a more versatile tool than the ones that existed before.

What research are you tackling now?

I have three directions related to CRISPR. The first one is to continue to explore the biochemistry of the CRISPR-Cas9 system. We also study the CRISPR-Cas9 system of other bacteria, besides S. pyogenes. We work on other aspects of CRISPR-Cas that are not related to the DNA targeting mechanism. I have some collaborations where we are developing the technology to deliver the CRISPR-Cas9 tool to certain types of human cells with the prospective to treat human genetic disorders. And I have founded a biotech company called CRISPR Therapeutics.

Are you studying other systems as well, or has CRISPR become the defining area of your work?

We are still working on other types of RNA-mediated regulatory mechanisms. For example, we are working on elucidating a regulatory mechanism involved in the degradation of proteins. We also study how cells communicate with one another and how the human host defends itself against bacteria.

I’m interested in working on CRISPR-Cas, but not exclusively. I want to spend the rest of my career doing basic science and trying to decipher new mechanisms that potentially could have some effect in medicine.

What do you see as the most exciting potential applications of CRISPR?

It has major implications for biotechnology and biomedicine. Companies have started to use it to form a library of knock-outs of human cells [cells with gene deletions] to screen for novel targets for new therapeutics. We can also generate animal models that are better for testing new therapeutics, because the model may mimic what is happening to humans more accurately.

I think with CRISPR-Cas9, human genetic disorders could be cured. Before we do so, a tool that enables delivery of this system to the cells must be developed. For genetic disorders that can be treated specifically using cell replacement, the technology could work.

What are your greatest concerns about how CRISPR might be used?

The biggest concern I have is misuse of this tool—for example, for the manipulation of human germlines or for purposes that would not be environmentally friendly. I’m not specifically against the manipulation of plants, but it depends on what is changed. There are still some years to go before the technology could work in humans. In Europe we have some ethical committees that have worked over the last 10 years on texts that forbid the manipulation of the genome of human germlines. Maybe in the United States it is not that fixed. There will certainly be some debates.

Chinese researchers recently published a paper showing that CRISPR-Cas9 could be used to edit human germlines. That work got a lot of media attention and was seen by some as a pivotal moment in the history of medicine. What was your reaction?

This particular article was accepted within one day after submission to the journal, which means that it had not been reviewed. I found this quite incredible.

This paper is a signal. Now some people have tried to edit human embryos, so maybe it is a good time to discuss the implications, because policy decisions always take a long time. Scientists together with developers, pharmaceutical industries, biotech, clinicians, experts in ethical issues, and the public need to reach a consensus on how the tool should be used. The public should not be afraid. There are good applications for this tool. It is important to educate everyone about what it is, what one can do with it, and how it works. We need to agree on certain usages that should be forbidden at the international level, including in Asian countries.

Research in genomic engineering is moving very quickly. What does the public need to know to keep up?

The first message I think people need is that this is a technology that originates from our doing pure, basic science. Politicians and funding agencies need to support basic science, because all discoveries, whether it’s in biology, chemistry, physics, or other fields, come from this kind of research.

The second message the public should understand is that it may be true that this tool could be misused, but it is also helping biologists at many different levels. There are still a number of genes with unknown functions. We are still far from understanding how molecules work and function together in a coordinated manner. This tool will help facilitate these genetic studies. The public is really afraid of the word transgenic, but CRISPR-Cas9 could be more precise than many current medicines, potentially with fewer secondary effects.

Do you think that there should be an international moratorium on editing germline cells, as was recently called for by Jennifer Doudna and others in a publication in Science?

Yes, surely, especially to avoid the types of publication we have seen lately [the Chinese paper].

Your work on CRISPR-Cas9 has triggered contentious patent issues. Is there a better way to handle them?

Yes, the patent issue is relatively complex. The technology can be widely used for many different applications, including human use, hence trying to secure some intellectual property is potentially hugely valuable. I think it will take some time to resolve these issues.

Despite this situation, it has not stopped anyone. I think this is good, because it could have had a detrimental effect on the development of the technology, and this has not been the case. What’s important is that everyone is using it, everyone wants to develop it, everyone has started to develop it, and no one has been blocked in the process.

You have founded a company, CRISPR Therapeutics, to commercialize your work. How did that come about?

Prior to CRISPR-Cas9, I was working on several molecular mechanisms involved in bacteria-caused diseases, and I was in contact with biotech companies that were focusing on the development of antibacterials and anti-infectives. So, I always had an eye on how a biotech company could be developed. But I knew one thing: If I wanted to found a new company, I needed to have an exciting finding that could be translated. With CRISPR-Cas9, very quickly, I saw that the tool indeed could be used for multiple purposes—as a tool for biotechnology and for biomedical purposes. I had this in mind, even before we showed how to obtain cleavage of DNA with this system.

I filed a patent application jointly with my collaborator Jennifer Doudna, but I was in Sweden, which might be the only country in the world where the inventor is still the owner of her or his own intellectual property. I knew people who were interested in collaborating with me. Very quickly, we found some investors.

I was lucky enough to have my intellectual property, so I have a little bit more power over what I wanted to do than some scientific founders. I put a lot of myself into this project. I wanted to make sure that I could see how it develops. It’s a little bit like when it’s your baby and you want to see the first steps and see where it goes.

Your CRISPR discovery has brought fame and big prizes. What have you learned from the experience?

My way of doing research was always a bit criticized because I did different things, but at the end of the day, for the CRISPR-Cas9 system it was good, because I could apply different expertise that I had acquired over the years and different types of thinking. It’s not so bad to have a multidisciplinary experience and work on different aspects of biology.

Sometimes I wonder whether I would have had my research funded if I had proposed exactly what I wanted to do—to decipher the CRISPR-Cas9 system. It’s an example that shows how important it is to provide scientists with some budget where they are allowed to do some blue-sky research. For sure one always needs to have some kind of big hypothesis or direction or interest, but sometimes you just hit something, certain components that you want to put together in a way that maybe does not make sense but you just want to do it. Scientists need to have the support to be able to do some crazy experiments to see where they go. My discovery has been a lesson for me that it’s important to give scientists the freedom and the time to do such work.