The CRISPR-cas9 technique enables researchers to precisely cut genes from DNA and paste a new piece of DNA to repair, for instance, a defective gene without further damaging the DNA code. ‘The molecule not only has a pair of scissors, but also a navigation system able to read DNA. As a result, it knows exactly where to go and where to cut the DNA’, Foijer says. ‘All genes have their own code, i.e. a unique letter combination. By providing CRISPR-cas9 with a gene’s barcode, it will look for the right location of the DNA. CRISPR-cas9 is highly innovative due to its scissors combined with its navigation system.’
Defence mechanism in bacteria
CRISPR-cas9 was not developed based on a theoretical concept, but discovered in bacteria at the late 1990s. It is a defence mechanism in bacteria used against viruses. Like humans, bacteria are constantly attacked by viruses. These are called bacteriophages. However, bacteria have a security system called CRISPR-cas9. This system is able to remember parts of genetic codes of viruses previously invading the bacteria and sets up a ‘library’ of genetic information in its own DNA. Bacteria use this CRISPR ‘genetic library’ to recognize these viruses when they re-attack the bacteria, and use the Cas9 enzyme to approache them and cut the viral DNA into pieces.
‘A host of scientists have contributed to this important preparatory study on bacteria, including John van der Oost, microbiologist at Wageningen University’, Foijer says. ‘Jennifer Doudna and Emmanuelle Charpentier, the two Nobel Prize winners, came up with the luminous idea to hijack this bacterial defence mechanism against viruses and develop it into an instrument suitable for use in human cells. That idea proved to be worth a Nobel Prize.’
CRISPR-cas9 and stem cell technology
The necessity to cleave DNA in order to change it is not a new insight. Since the 1990s, scientists have been using a technique aimed at making adjustments to, for instance, mouse cells. ‘Although this old technique allowed cell changes to be made, it proved to be a very inefficient method that could not be used for human cells, for instance’, Foijer says. ‘We could not exactly target the break site of the DNA, which is required to make changes to the DNA code. Instead, we relied on spontaneous breaks in the DNA nearby the gene that we wanted to edit, so the desired change only occurred in very few cells or not at all.'
The development of CRISPR-cas9 in 2013 proved to be the required ‘sniper’ required to precision-edit the DNA of human cells. Researchers around the world have been eager to use it ever since. Foijer combines CRISPR-cas9 with stem cell technology: ‘Stem cell technology enables us to make all kinds of tissues for research purposes. This provides us with a great deal of information about hereditary diseases. We can use CRISPR-cas9 to repair defective genes in stem cells and to make new tissues from it. Eventually, we may be able to transplant these tissues.’
Another important application of CRISPR-cas9 is genetic screens. Recently, Foijer’s research group was awarded a grant for this by KWF Dutch Cancer Society. One example of such a screen is a drug-resistance screen. ‘Although we do not know why, cancer cells may become resistant to chemotherapy’, Foijer says. ‘In such a genetic screen we grow millions of cancer cells, after which a host of gene-specific Cas9 proteins are released, each cutting a different gene in individual cancer cells. Next, a chemotherapeutic agent is added. After a while, only the drug-resistant cells remain. We next determine which genes in these cells have been eliminated by CRISPR-cas9. This, reveals which genes are important to protect against resistance for the tested drug and thus enables us to gain more knowledge about the resistance mechanism.’
Turning off genes
‘In theory, cas9 enables us to do everything we would like to do with DNA’, Foijer says. ‘Most importantly, the Cas9 protein can be sent to a specific location. Engineering other protein elements onto the Cas9 protein can be used to change its function. For instance, one can remove the 'scissors' and make Cas9, instead of cutting the DNA, turn on or off a certain gene without changing the code. This is particularly relevant as some diseases are not caused by a mutation, but by a gene being turned on or off. Since the discovery of CRISPR-cas9, new applications have been found each month. The possibilities are endless’, the researcher says.