Bence Kover introduces how the molecular scissor CRISPR can be used for unconventional purposes such as diagnostics, providing a simple and cheap testing method for COVID-19.
Designer babies, superhuman armies, and a utopian society without disease and aging – these are the things that laymen commonly, but quite wrongfully, associate CRISPR with. While CRISPR certainly has nothing to do with all the above, it is unquestionably going to be a defining technology of the coming decade.
On 7 October 2020, Jennifer Doudna and Emmanuelle Charpentier received the Nobel Prize in Chemistry for their work on CRISPR gene editing. Together with all the breakthroughs of genomics, such as cheap genetic sequencing, CRISPR has been promised to revolutionize medicine, agriculture, and life sciences research. One of the unforeseen applications of CRISPR however, is in the field of diagnostics. Research in this field has advanced rapidly, largely catalysed by the increasing demand for nucleic acid detection technologies during the coronavirus pandemic. Will this technology change COVID testing, and pave the way for CRISPR’s future?
At the heart of CRISPR lies a protein, called a restriction enzyme, that enables the cleavage of DNA. In genetic engineering, they use the restriction enzyme CAS9 (CAS = CRISPR Associated). This enzyme makes a precise cut in the DNA and subsequently gets inactivated, meaning that the job of CAS9 is over. Genetic engineering relies on the cell’s own ability to fix this break in the DNA, which commonly results in an introduced mutation, or allows the researchers to insert a gene. There are however other CRISPR restriction enzymes that behave slightly differently. The protein Cas12, for example, also makes a precise cut at a specific location in the DNA, however it then remains activated and goes on to cleave any single stranded DNA. Normally this would not be apparent as DNA tends to be double stranded, hence the name double helix. However, if we supply single stranded DNA with fluorescent labels, let’s call them “reporters”, to the reaction mixture, they will get cleaved by Cas12. To summarise, if Cas12 recognises its initial target sequence it will get activated, and signal this to us by cleaving the fluorescently labelled reporters, therefore also signalling the presence of its target DNA.
Based on this idea, the technology called DNA Endonuclease-Targeted CRISPR Trans Reporter, or DETECTR for short, was born. In theory, any kind of nucleic acid sample, RNA or DNA, coming from a virus, bacteria or any higher organism can be detected with this method. However, because the activity of Cas12 only applies to DNA, any RNA sample has to first be converted to DNA. This is notably true for SARS-CoV-2, the virus causing COVID-19, which is a single stranded RNA virus and is the number one target of DETECTR currently.
A SARS-CoV-2 DETECTR test takes around 40-60 minutes, which is considerably lower than the 4-6 hours a PCR test would take, where the sampling to diagnosis time is usually more than 24 hours. The test also seems to provide reliable results, as it has been shown to have 95% sensitivity and 100% specificity. Besides the rapid and precise diagnosis, this method also requires much simpler equipment than the current gold-standard PCR, which necessitates both complex equipment and a professional laboratory environment.
DETECTR has already been shown to be effective in diagnosing SARS-CoV-2, and other viruses, such as human papillomavirus HPV16 and HPV18. On October 29th, Mammoth Biosciences, co-founded by Doudna, announced a partnership with the pharma giant Merck, to cooperate in large-scale production of DETECTR tests. By the end of 2020 they are looking to receive an FDA Emergency Use Authorization due to the current pandemic. They claim that their DETECTR-based machines will be “high-throughput systems [that] will be compatible with both nasal swab and saliva samples and are targeting 1500 tests per 8-hour shift with minimal user interaction”. Mammoth Biosciences had a series B fundraising in early 2020 where they raised 45 million USD from investors and is currently one of the leaders of the CRISPR industry.
The field of CRISPR research is relatively polarized, and behind every discovery or launched company, there is generally one of two groups: the camp of the now Nobel laureate Doudna or the Chinese-American Feng Zhang. While the DETECTR method was worked out by Doudna’s team, a similar method called Specific High-sensitivity Enzymatic Reporter unLOCKing, or SHERLOCK for short, was already being worked on by Zhang’s lab. SHERLOCK is based on a different CAS protein, called Cas13, which has a cleavage activity on single-stranded RNA instead of single-stranded DNA. SHERLOCK has been used for the detection of Dengue, Zika, Malaria and, of course, SARS-CoV-2. The SHERLOCK technology has already received an FDA emergency use authorization, and SHERLOCK Biosciences, co-founded by Zhang, has announced a partnership in June with binx health to scale up the production of their test. While these tests will hopefully be available for laboratories by 2021, the ultimate goal of SHERLOCK Biosciences is to bring testing to the homes of people.
While technologies promising to revolutionize diagnostics are faced with doubt since the 2016 Theranos scandal, CRISPR based diagnostics do have a potential that is hard not to recognize. Hopefully this technology will have a large enough positive impact on the public and the investors, to counter-balance the resistance against gene-therapies and GMO crops, and pave the way for the “true” applications of CRISPR.
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