CRISPR-Cas Inhibiting Polypeptides

Technology Description

The inventions introduce novel Cas nuclease inhibiting polypeptides with synthetic/ non-natural amino acids as well as techniques to conjugate it to an antibody or PEG for easy delivery to targeted cells. Bacterial cells respond to phage attack by activating CRISPR-Cas nuclease to digest phage genome; to escape it, phages encode anti-CRIPSR-Cas proteins. Broad-spectrum Cas enzyme inhibitors will allow for better post-translational regulation of CRISPR-Cas system for gene editing. These polypeptides were hitherto unknown and their use along with CRISPR-Cas system for genetic manipulation will improve its efficacy and potential for translation to human applications. 

Value Proposition

• Anti-CRISPR polypeptides are the tools which will efficiently target the genes being edited
• Large number of clinical trials are currently using CRISPR-Cas systems to treat diseases- from hematological malignancies to treating HIV via permanent genetic modifications.  However, a major risk involved in using CRISPR-Cas based gene therapy are off-target effects, which may lead to permanent changes in patient genome at non-target sites. 
• Anti-CRISPR polypeptides allow for long-term regulation of CRISPR-Cas therapy without any off-target effects. They can be introduced independently of the CRISPR system, and they will maintain activity over longer durations.
• Anti-CRISPR polypeptides act with a better specificity are easier to design and synthesize, when compared to small molecule inhibitors.

Related Materials

• Marino, N. D., Pinilla-Redondo, R., & Bondy-Denomy, J. (2022). CRISPR-Cas12a targeting of ssDNA plays no detectable role in immunity. BioRxiv, 2022.03.10.483831.
• Rauch BJ, Silvis MR, Hultquist JF, Waters CS, McGregor MJ, Krogan NJ, Bondy-Denomy J. Inhibition of CRISPR-Cas9 with Bacteriophage Proteins. Cell. 2017 Jan 12;168(1-2):150-158.e10.
• Mahendra C, Christie KA, Osuna BA, Pinilla-Redondo R, Kleinstiver BP, Bondy-Denomy J. Broad-spectrum anti-CRISPR proteins facilitate horizontal gene transfer [published correction appears in Nat Microbiol. 2020 Jun;5(6):872]. Nat Microbiol. 2020;5(4):620-629.

Patent Status

Additional Patents Pending