At the forefront of biological advances in the 21st century is the CRISPR-Cas9 genome editing system, which employs “Clustered Regularly Interspaced Short Palindromic Repeats” present in DNA sequences and the Cas9 endonuclease protein to edit genomic sequences. This system was first observed in a number of bacterial species as a defense against viral infection, in which viral DNA is assimilated into the host’s genomic CRISPR sequence.2 This is then transcribed into CRISPR RNA that Cas9 recognizes and degrades, resulting in the destruction of the introduced foreign genome. In addition to providing an effective and adaptive immune system in prokaryotic organisms, exogenous CRISPR-Cas9 from prokaryotes introduced to eukaryotes has been shown to precisely execute double stranded cuts in DNA sequences. This is accomplished by introducing a complementary DNA-specific strand of guide-RNA (gRNA) that binds to the target DNA sequence, which is then subsequently recognized by Cas9. Following recognition, the gRNA-DNA complex is cut out of the DNA sequence and the two ends are ligated together, allowing for the complete deletion of a targeted gene. Alternatively, an additional DNA sequence can be introduced into the cut, allowing for the manipulation of the genetic sequence.
The beauty of the CRISPR-Cas9 system is in its cost-effective and precise editing of DNA sequences. If researchers have access to the nucleotide sequence of a gene of interest, they can cheaply and quickly synthesize complementary gRNA, which will target the gene for digestion by Cas9, resulting in either the deletion or replacement of that gene. In a study conducted by James DiCarlo of Columbia University using the yeast species Saccharomyces cerevisiae, “co-transformation of a gRNA plasmid and a donor DNA in cells constitutively expressing Cas9 resulted in near 100% donor DNA recombination frequency.”2 The system’s accuracy demonstrates its potential to phase out alternative methods of modifying gene expression and advance every aspect of the biological sciences.
If the efficacy and accuracy the system displayed in the manipulation of simpler organisms carries over to the study of complex eukaryotes, CRISPR-Cas9 could allow for biologists to engineer “perfect” organisms, devoid of malign genetic characteristics and teeming with preferential genes.7 Due to the potential of the CRISPR Targeted Gene Editing System, the rights to the requisite technology have been hotly contested, primarily between the Feng Zhang lab of the Broad Institute, based out of Harvard University and the Massachusetts Institute of Technology, and Jennifer Doudna and Emmanuelle Charpentier’s team at the University of California at Berkeley. While these two research teams are by no means the only ones to have achieved remarkable breakthroughs in the discovery and harnessing of CRISPR-Cas9, they are at the vanguard of the legal battle, because their respective work is essential for genomic editing by the system. In 2012, Charpentier and Doudna characterized the biochemical nature of Cas9 mediated double-stranded DNA cleavage and discovered simpler mechanisms for the direction of gRNA toward target DNA sequences,3 and applied this technology to alter prokaryotic DNA.4 Subsequently, in May of 2012, Doudna filed a patent claim for the molecular machinery the team had developed. In December of 2012, however, Zhang had harnessed the capability of the system to successfully edit the genomic sequences of eukaryotic organisms.5 He subsequently filed an expedited patent for his adaptation of the system in eukaryotes and was granted the rights to the system in eukaryotes in 2014. This prompted UC Berkeley to engage in “patent interference”, a legal maneuver in which the university requested the United States Patent and Trademark Office to determine the original developer of the system.
On February 15th, 2017, after over two years of heated legal dispute, the United States Patent and Trademark Office’s Federal Patent Trial and Review Board ruled that the CRISPR-Cas9 technology employed by Feng Zhang and the Broad Institute to modify eukaryotic cells differed significantly compared to the technology employed by Doudna’s team to modify prokaryotic cells.6 The board thus determined that the Broad Institute’s claim to the rights for the system of editing genes in eukaryotic organisms does not encroach upon UC Berkeley’s claim to the rights for the system of genomic editing in prokaryotes, even though Berkeley had filed a patent claim for the general CRISPR-Cas9 system in 2012, months before the Broad Institute had filed claims for the system of eukaryotic genomic editing. UC Berkeley has filed an appeal, arguing that the two systems, prokaryotic and eukaryotic, are inseparable based on the molecular machinery involved, and thus the original discoverers, Doudna and Charpentier, lay claim to the patent for the use of the system in both prokaryotes and eukaryotes. However, the Broad Institute’s legal team has cited Doudna as saying that she “had trouble adapting CRISPR-Cas9 in eukaryotic cells,” which the court considers evidence that the adaptation of the system to eukaryotic cells was independently developed by the efforts of the research team at the Broad Institute.
While the dispute is not over yet, resolution has never been closer, and the Broad Institute appears to be on the cusp of possessing the rights to one of the most significant biotechnological advancements of the century. Not only would the Broad Institute and its associated investors incur gargantuan profits, but it would also control the ethics behind furthering the technology and application of the system in the most complex organisms, humans. Since 2016, Lu You at Sichuan University in China has been introducing the CRISPR-Cas9 system to attack malignant proteins in lung cancer patients.1 While You has not published any results of the study, he has certainly incited a firestorm of curiosity and criticism over the use of the system in humans. The nature of the system could allow for the elimination of detrimental genes from individuals, as well as the substitution of functional counterparts. But if the system could be used to amend genetic defects, it could also be used to change any number of traits, from eye color to height. The implications of engineering human beings via CRISPR-Cas9 are revolutionary and definitely something to keep an eye on for the future.
- Cyranoski, David. “CRISPR gene-editing tested in a person for the first time.” Nature. 15
- DiCarlo, JE., et al. “Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas
Systems.” Nucleic Acids Res. 04 March 2014.
- Doudna, JA., Charpentier, E. “Genome Editing: The New Frontier of Genome Editing with
CRISPR-Cas9.” Science. 28 November 2014.
- Doudna, JA. “The Doudna Lab: Exploring Molecular Mechanisms of RNA-Mediated Gene
Regulation.” Web. Accessed 20 February 2017. http://rna.berkeley.edu/crispr.html
- Makarova KS., Zang F., Koonin, EV. “SnapShot:Class 2 Crispr-Cas Systems.” Cell. 12
- Martin, Dermot. “Ruling in CRISPR-Cas9 Patent Battle.” Web. Accessed 20 February 2017.
- Reardon, Sara. “CRISPR heavyweights battle in US patent court.” Nature. 06 December