Unmet Need
Protein cleavage is a fundamental molecular process, essential to cellular viability and tissue physiology across all kingdoms of life. Human proteases comprise a diverse set of enzymes with at least 569 members, making up five distinct catalytic classes. Cancer associated protease activities are often dysregulated and usually contribute to the selective growth advantage and metastatic potentialof tumor cells. There is thus a need for a comprehensive assay to characterize protease activities within biological samples.
Technology Overview
Researchers have found that detection technologies based on DNA analysis can provide extremely high sensitivity, and have used high throughput DNA synthesis to encode complete proteomes. Each of about 30,000 humanprotein isoforms were individually tiled using 90-aa peptides with 45-aa overlaps. The peptide-encoding DNA sequences were synthesized and cloned into a novel T7 Sensing EndoPeptidase Activities via Removal of Tag Epitopes (SEPARATE) phage display system. One peptide molecule per phage particle is displayed, such that the peptides are flanked by an N-terminal FLAG tag and C-terminal biotinylation. The biotin-immobilized library is digested and cleavage-released phage are re-captured on beads. The uncleaved and recaptured phage populations are then sequenced for unbiased, quantitative detection of all peptide cleavage events.
Stage of Development
Prototypes have been completed and are in the testing phase.
Patent Information
PROTEASE ACTIVITY PROFILING VIA PROGRAMMABLE PHAGE DISPLAY OF COMPREHENSIVE PROTEOME-SCALE PEPTIDE LIBRARIES Pending United States application: 16/913,572
Filed: 6/26/2020
https://patents.google.com/patent/US20200407467A1/en?oq=16%2f913%2c572
Publication
Gabriel D. Román-Meléndez, Thiagarajan Venkataraman, Daniel R. Monaco, H. Benjamin Larman,
Protease Activity Profiling via Programmable Phage Display of Comprehensive Proteome-Scale Peptide Libraries,
Cell Systems, Volume 11, Issue 4, 2020, Pages 375-381.e4, ISSN 2405-4712
https://doi.org/10.1016/j.cels.2020.08.013.
(https://www.sciencedirect.com/science/article/pii/S2405471220303240)
