Our goal is to develop a transition metal agent that specifically inhibits matrix metalloproteinase (MMP)-2 by a unique mechanism. Such an agent may have potential use in antimetastatic therapy. A MMP-2 specific peptide substrate will be tethered to a Co(III)-Schiff base (Co(III)-sb) complex that has been previously shown to irreversibly inhibit histidine (His)-containing enzymes. The MMP-2 targeting peptide will impart specificity for MMP-2 over other enzymes and bring the Co(III)-sb complex to close proximity of the target protein such that selective inhibition of MMP-2 occurs. The peptide conjugate is expected to show low levels of off-target effects while maintaining effective inhibition of the target enzyme. Peptides known to be selectively cleaved by MMP-2 over other MMPs will be synthesized and tethered to Co(III)-sb (Co(III)-pep), and fluorescent analogues (Co(III)-flupep) will be synthesized to allow monitoring of their action and localization in biological systems usin fluorescence microscopy. The effectiveness of Co(III)-pep and Co(III)-flupep as a MMP-2-specific inhibitor will be evaluated by investigating kinetic and thermodynamic interactions between the inhibitor and enzyme. In addition, the specificity of Co(III)-pep and Co(III)-flupep fo MMP-2 over other enzymes will be investigated by screening them against a range of MMPs and non-MMP enzymes using inhibition assays and zymography. The binding and inhibitory activity of Co(III)-pep and Co(III)-flupep will be compared to the non-targeted Co(III)-sb complex to evaluate the suitability of targeted Co(III) complexes as MMP-2 specific inhibitors. Further, the effect of Co(III)-pep on the invasion and migration of various cancer cell lines will be validated by testing in biological systems. Migration and invasion assays on cell monolayers and 3-dimensional tumor models such as spheroids will be carried out to evaluate the effect of the peptide conjugates in inhibiting metastatic activity of the cell. Immunofluorescence assays will be performed to ascertain the extent of MMP-2 inhibition in spheroid sections. Upon completion of the proposed in vitro biological studies and if successful results are obtained, we believe this novel inhibitor can be utilized as an antimetastatic agent in vivo.

Public Health Relevance

Our goal is to develop a specific inhibitor of matrix metalloproteinase (MMP)-2 as a potential antimetastatic agent with a unique mechanism of action. A peptide substrate that will impart specificity for MMP-2 over other enzymes will be tethered to an irreversible cobalt(III) enzyme inhibitor to bring it in close proximity of the targt. This approach will decrease off-target effects while maintaining effective inhibition of the target enzyme, giving rise to an efficient and viable inhibitor of MMP-2.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Small Research Grants (R03)
Project #
1R03CA167715-01A1
Application #
8512054
Study Section
Special Emphasis Panel (ZCA1-SRLB-2 (J1))
Program Officer
Perloff, Marjorie
Project Start
2013-04-01
Project End
2015-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
1
Fiscal Year
2013
Total Cost
$77,250
Indirect Cost
$27,250
Name
Northwestern University at Chicago
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
160079455
City
Evanston
State
IL
Country
United States
Zip Code
60201
Heffern, Marie C; Velasco, Pauline T; Matosziuk, Lauren M et al. (2014) Modulation of amyloid-? aggregation by histidine-coordinating Cobalt(III) Schiff base complexes. Chembiochem 15:1584-9
Peterson, Mark D; Holbrook, Robert J; Meade, Thomas J et al. (2013) Photoinduced electron transfer from PbS quantum dots to cobalt(III) Schiff base complexes: light activation of a protein inhibitor. J Am Chem Soc 135:13162-7