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Evaluation of a triple-helical peptide with quenched fluorophores for optical imaging of MMP-2 and MMP-9 proteolytic activity

Zhang, X and Bresee, J and Cheney, PP and Xu, B and Bhowmick, M and Cudic, M and Fields, GB and Edwards, WB (2014) Evaluation of a triple-helical peptide with quenched fluorophores for optical imaging of MMP-2 and MMP-9 proteolytic activity. Molecules, 19 (6). 8571 - 8588.

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Abstract

Matrix metalloproteinases (MMP) 2 and 9, the gelatinases, have consistently been associated with tumor progression. The development of gelatinase-specific probes will be critical for identifying in vivo gelatinoic activity to understand the molecular role of the gelatinases in tumor development. Recently, a self-assembling homotrimeric triple-helical peptide (THP), incorporating a sequence from type V collagen, with high substrate specificity to the gelatinases has been developed. To determine whether this THP would be suitable for imaging protease activity, 5-carboxyfluorescein (5FAM) was conjugated, resulting in 5FAM3-THP and 5FAM6-THP, which were quenched up to 50%. 5FAM6-THP hydrolysis by MMP-2 and MMP-9 displayed kcat/KM values of 1.5 × 104 and 5.4 × 103 M -1 s-1, respectively. Additionally 5FAM6-THP visualized gelatinase activity in gelatinase positive HT-1080 cells, but not in gelatinase negative MCF-7 cells. Furthermore, the fluorescence in the HT-1080 cells was greatly attenuated by the addition of a MMP-2 and MMP-9 inhibitor, SB-3CT, indicating that the observed fluorescence release was mediated by gelatinase proteolysis and not non-specific proteolysis of the THPs. These results demonstrate that THPs fully substituted with fluorophores maintain their substrate specificity to the gelatinases in human cancer cells and may be useful in in vivo molecular imaging of gelatinase activity. © 2014 by the authors.


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Details

Item Type: Article
Status: Published
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Zhang, X
Bresee, J
Cheney, PP
Xu, B
Bhowmick, M
Cudic, M
Fields, GB
Edwards, WB
Date: 1 January 2014
Date Type: Publication
Journal or Publication Title: Molecules
Volume: 19
Number: 6
Page Range: 8571 - 8588
DOI or Unique Handle: 10.3390/molecules19068571
Schools and Programs: School of Medicine > Radiology
Refereed: Yes
Date Deposited: 12 May 2015 18:28
Last Modified: 05 Feb 2019 17:55
URI: http://d-scholarship.pitt.edu/id/eprint/24839

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