Abstract
Cashew Nut Shell Liquid (CNSL) has been used in traditional medicine for the treatment of a wide variety of pathophysiological conditions. To further define the mechanism of CNSL action, we investigated extracts of CNSL (CNSE) on two matrix metalloproteinases, MMP-2/Gelatinase-A and MMP-9/Gelatinase-B that are known to have critical roles in several disease states. We observed that the major constituent of CNSE, anacardic acid, markedly inhibited the gelatinase activity of 3T3-L1 cells. Our gelatin zymography studies on these two secreted gelatinases, present in the conditioned media from 3T3-L1 cells, established that anacardic acid directly inhibited the catalytic activities of both MMP-2/9. Our docking studies suggested that anacardic acid binds into the MMP-2/9 active site, with the carboxylate group of anacardic acid chelating the catalytic zinc ion, and forming a hydrogen bond to a key catalytic glutamate side chain, and the C15 aliphatic group being accommodated within the relatively large S1′ pocket of these gelatinases. In agreement with the docking results, our fluorescence-based studies on recombinant MMP-2 catalytic core domain demonstrated that anacardic acid directly inhibits substrate peptide cleavage in a dose-dependent manner, with an IC50 of 11.11μ M. Additionally, our gelatinase zymography and fluorescence data confirmed that cardol-cardanol mixture, salicylic acid and aspirin, all of which lack key functional groups present in anacardic acid, are much weaker MMP-2/-9 inhibitors. Our results provide the first evidence for inhibition of gelatinase catalytic activity by anacardic acid, providing a novel template for drug discovery and a molecular mechanism potentially involved in CNSL therapeutic action.
- Structure-activity relationships and modeling
- Fluorescence techniques
- Mass Spectroscopy
- NMR
- Protein-binding
- Enzymology
- Structure/function/mechanism
- Matrix metalloproteins (MMPs)
- Received March 29, 2012.
- Revision received June 28, 2012.
- Accepted June 28, 2012.
- The American Society for Pharmacology and Experimental Therapeutics