Gene editing is still mostly used for research, but healthcare market research firm Kalorama Information says there are three reasons to think that diagnostics applications for CRISPR are well on their way. One is the improvement of a RNA guided editing technique.
ROCKVILLE, Md., April 24, 2018 /PRNewswire/ -- Gene editing is still mostly used for research, but healthcare market research firm Kalorama Information says there are three reasons to think that diagnostics applications for CRISPR are well on their way. One is the improvement of a RNA guided editing technique. Then there is a new diagnostic technique, and an important retraction of criticism that was beginning to lead many to question the entire technology.
“The CRISPR Diagnostic toolkit has gotten handier,” said Kalorama analyst Rob Camp, PhD, in a recent blog post.
One: A New Version of SHERLOCK: SHERLOCK--Specific High-sensitivity Enzymatic Reporter unlocking--is derived from CRISPR/Cas; like its progenitor, the technique employs guide RNAs paired up with enzymes, seeking out specific DNA sequences and cutting them. However, the Cas13a enzyme used in SHERLOCK (as opposed to CRISPR’s Cas9) makes additional cuts near the target cleavage site, as well as to an additional reporter RNA molecule that releases a fluorescent signal when cleaved. The method showed vast potential as an in vitro diagnostic technology. Now the technique is back in the news with an update. As reported in an article published by the AAAS on 15 February, SHERLOCKv2 shows a 3.5-fold increase in sensitivity over its original iteration; responsible for this is the use of Csm6, an auxiliary CRISPR-associated enzyme, in addition to Cas13. It was found that not only does this further amplify the signal, but it provides for more rapid detection by lateral flow, making the technique even more viable for applications in paper-based diagnostic tests without the need for expensive, complex machinery. Furthermore, researchers, including original CRISPR developer Jennifer Doudna, were able to use SHERLOCK for multiplex detection, reporting that they designed a test able to simultaneously find Zika and Dengue virus ssRNA as well as a synthetic ssRNA target. With preamplification by recombinase polymerase amplification, single-molecule detection can be achieved at the attomolar level.
Two: DETECTER -Described in another article published on the same day is an altogether new platform derived from CRISPR/Cas. A team led by another CRISPR pioneer, Feng Zhang, reports that the enzyme Cas12a, after binding to and cleaving the target DNA sequence, will continue to cleave and ultimately completely degrade surrounding ssDNA molecules. Combining this ssDNase activation with isothermal amplification, Zhang and crew developed a technique they call DNA Endonuclease Targeted CRISPRTransReporter, or DETECTR, which, similarly to SHERLOCK, can achieve attomolar sensitivity for DNA detection. In the study, DETECTR was used to specifically detect human papillomavirus strains 16 and 18 from cultured human cells (SiHa, HeLa, and uninfected BJAB as control) as well as from patient anal swab samples within one hour, as a proof-of-concept for a new molecular diagnostic platform.
Three: Pushback on CRISPR Criticism: Finally, in unrelated-but-still-somewhat-related news: In the previous blog entry on CRISPR, we briefly mentioned that potential issues could emerge in the form of mosaics produced by the guide RNA missing target sequences, resulting in widespread damage to the genome. However, this week, Nature Methods has retracted an article published in May 2017 outlining the danger. Initially, the article sparked an outcry from a number of people, some of whom were associated with companies with licensing rights to CRISPR; criticism included assertions that the investigators did not use proper controls to ensure that mutations were not results from normal variation in subjects. Nature Methods itself published an editorial statement of concern in July 2017, and issued the formal retraction in late March. The authors, led by Stanford’s Vinit B Mahajan, acknowledged that the criticism may be on point; Mahajan and three coauthors nonetheless disapproved of the retraction, however, stating that the matter requires further study before such a statement can be made in full faith. Follow-up studies are underway using whole-genome sequencing.
The firm pegged CRISPR-related technology sales at $779 million in 2017 with fast revenue growth expected. For more information on CRISPR and the associated businesses see Kalorama’s white paper on gene editing and full report on the technique’s markets.
About Kalorama Information
Kalorama Information, a division of MarketResearch.com, supplies the latest in independent medical market research in diagnostics, biotech, pharmaceuticals, medical devices and healthcare; as well as a full range of custom research services. Reports can be purchased through Kalorama’s website and are also available on www.marketresearch.com and www.profound.com.
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