Unmet Need
Since CRISPR has been proposed as a genome editing tool in 2012, it has revolutionized the field and created excitement extending beyond the genome editing scientific community. Due to its simplicity and efficiency, CRISPR-Cas9 systems have already shown promise in multiple fields, e.g., medicine, agriculture and even biofuels (through CRISPR-edited algae).
CRISPR systems are evolving rapidly with increasing understanding of its mechanism of action(s), novel applications and innovative modifications. One important example of CRISPR modification is chemically-induced initiation of Cas9 cutting activity. While chemical induction is effective, it results in slow kinetics of CRISPR activity and requires the nuclear accumulation of active Cas9 after induction. This delays its overall efficiency and hinders the ability for tight control of CRISPR/Cas9 activity. As the market for biomedical engineering, and in particular, genome editing, continues to grow, it is necessary to develop faster, better and more accurate CRISPR-based technologies.
Technology Overview
Johns Hopkins scientists have developed a very rapid and efficient CRIPSR/Cas9 system utilizing light induced rather than chemically induced Cas9 activation. In their design, light-sensitive caged nucleotides are strategically placed to create artificial mismatches as a “roadblock”. Upon UV light stimulation at 365 or 405nm, this roadblock is removed and the guide RNA can fully hybridize with the target DNA. Within 30 seconds of stimulation, cleavage of more than 50% of genomic DNA can be observed and indel formations reach ~90% within 6 hours, thereby demonstrating rapid and high efficiency of DNA cleavage and gene knockout. Additionally, Cas9 can be activated rapidly in single cells and/or locally within cells around a targeted genetic focus. Using fluorescence imaging and a focused laser, this technology can achieve extreme spatial control of gene manipulation which is unmatched by any current CRISPR technology.
Stage of Development
This technology has been fully developed and validated for research and development. Additional work is planned to validate safety in potential therapeutic settings.
Publications
Zou RS, Marin-Gonzalez A, Liu Y, Liu HB, Shen L, Dveirin RK, Luo JXJ, Kalhor R, Ha T. Massively parallel genomic perturbations with multi-target CRISPR interrogates Cas9 activity and DNA repair at endogenous sites. Nat Cell Biol. 2022 Sep;24(9):1433-1444.
