Yonghao Jin

Matrix Metalloproteinase Inhibitors as Prospective Agents for the Prevention and Treatment of Cardiovascular and Neoplastic Diseases

Acting on a broad spectrum of extracellular, intracellular, and membrane-associated substrates, the matrix metalloproteinases (MMPs) are critical to the biological processes of organisms; when aberrantly expressed, many pathological conditions may be born or exacerbated. The prospect of MMP inhibition for therapeutic benefit in cancer, cardiovascular disease, and stroke is reviewed here. MMP inhibitor (MMPI) development constitutes an important branch of research in both academic and industrial settings and advances our knowledge on the structure-function relationship of MMPs. Targeting MMPs in disease treatment is complicated by the fact that MMPs are indispensable for normal development and physiology and by their multi-functionality, possible functional redundancy or contradiction, and context-dependent expression and activity. This complexity was revealed by previous efforts to inhibit MMP activity in the treatment of cancer patients that yielded unsatisfactory results. This review focuses on MMPI development since the late 90s, in terms of natural products and their derivatives, and synthetic compounds of low molecular mass incorporating specific zinc-binding groups (ZBGs). A few polyphenols and flavonoids that exhibit MMPI activities may have chemopreventive and neuro- and cardiovascular-protective effects. A new generation of potent and selective MMPIs with novel ZBGs and inhibition mechanisms have been designed, synthesized, and tested. Although only one collagenase inhibitor (Periostat, doxycycline hyclate) has been approved by the Food and Drug Administration as a drug for the treatment of periodontal disease, new hope is emerging in the form of natural and synthetic MMPIs for the prevention and treatment of stroke, cardiovascular disease, cancer, and other medical conditions.

The intermediate S1' pocket of the endometase/matrilysin-2 active site revealed by enzyme inhibition kinetic studies, protein sequence analyses, and homology modeling.

Human matrix metalloproteinase-26 (MMP-26/endometase/matrilysin-2) is a newly identified MMP and its structure has not been reported. The enzyme active site S1′ pocket in MMPs is a well defined substrate P1′ amino acid residue-binding site with variable depth. To explore MMP-26 active site structure-activity, a series of new potent mercaptosulfide MMP inhibitors (MMPIs) with Leu or homophenylalanine (Homophe) side chains at the P1′ site were selected. The Homephe side chain is designed to probe deep S1′ pocket MMPs. These inhibitors were tested against MMP-26 and several MMPs with known x-ray crystal structures to distinguish shallow, intermediate, and deep S1′ pocket characteristics. MMP-26 has an inhibition profile most similar to those of MMPs with intermediate S1′ pockets. Investigations with hydroxamate MMPIs, including those designed for deep pocket MMPs, also indicated the presence of an intermediate pocket. Protein sequence analysis and homology modeling further verified that MMP-26 has an intermediate S1′ pocket formed by Leu-204, His-208, and Tyr-230. Moreover, residue 233 may influence the depth of an MMP S1′ pocket. The residue at the equivalent position of MMP-26 residue 233 is hydrophilic in intermediate-pocket MMPs (e.g. MMP-2, -8, and -9) and hydrophobic in deep-pocket MMPs (e.g. MMP-3, -12, and -14). MMP-26 contains a His-233 that renders the S1′ pocket to an intermediate size. This study suggests that MMPIs, protein sequence analyses, and molecular modeling are useful tools to understand structure-activity relationships and provides new insight for rational inhibitor design that may distinguish MMPs with deep versus intermediate S1′ pockets.

Catalytic- and ecto-domains of membrane type 1-matrix metalloproteinase have similar inhibition profiles but distinct endopeptidase activities

Membrane type 1-matrix metalloproteinase (MT1-MMP/MMP-14) is a major collagenolytic enzyme that plays a vital role in development and morphogenesis. To elucidate further the structure-function relationship between the human MT1-MMP active site and the influence of the haemopexin domain on catalysis, substrate specificity and inhibition kinetics of the cdMT1-MMP (catalytic domain of MT1-MMP) and the ecto domain DeltaTM-MT1-MMP (transmembrane-domain-deleted MT1-MMP) were compared. For substrate 1 [Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH(2), where Mca stands for (7-methoxycoumarin-4-yl)acetyl- and Dpa for N -3-(2,4-dinitrophenyl)-L-2,3-diaminopropionyl], the activation energy E (a) was determined to be 11.2 and 12.2 kcal/mol (1 cal=4.184 J) for cdMT1-MMP and DeltaTM-MT1-MMP respectively, which is consistent with k (cat)/ K (M) values of 7.37 and 1.46x10(4) M(-1).s(-1). The k (cat)/ K (M) values for a series of similar single-stranded peptide substrates were determined and found to correlate with a slope of 0.17 for the two enzyme forms. A triple-helical peptide substrate was predicted to have a k (cat)/ K (M) of 0.87x10(4) M(-1).s(-1) for DeltaTM-MT1-MMP based on the value for cdMT1-MMP of 5.12x10(4) M(-1).s(-1); however, the actual value was determined to be 2.5-fold higher, i.e. 2.18x10(4) M(-1).s(-1). These results suggest that cdMT1-MMP is catalytically more efficient towards small peptide substrates than DeltaTM-MT1-MMP and the haemopexin domain of MT1-MMP facilitates the hydrolysis of triple-helical substrates. Diastereomeric inhibitor pairs were utilized to probe further binding similarities at the active site. Ratios of K (i) values for the inhibitor pairs were found to correlate between the enzyme forms with a slope of 1.03, suggesting that the haemopexin domain does not significantly modify the enzyme active-site structure.

Enhancing QM/MM molecular dynamics sampling in explicit environments via an orthogonal-space-random-walk-based strategy.

Accurate prediction of molecular conformations in explicit environments, such as aqueous solution and protein interiors, can facilitate our understanding of various molecular recognition processes. Most computational approaches are limited as a result of their compromised choices between the underlying energy model and the sampling length. Taking advantage of a recent second-order generalized ensemble scheme [e.g., the orthogonal space random walk (OSRW) strategy], which can synergistically accelerate the motion of a focused region and its coupled environmental response, we are presenting a QM/MM (combined quantum mechanical/molecular mechanical)-based molecular dynamics sampling technique to explore molecular conformational landscapes in explicit environments. The present QM/MM potential scaling-based OSRW sampling scheme is employed to study the binding of DMSO to the FKBP12 protein, the conformation distribution of a novel mercaptosulfonamide inhibitor in aqueous solution, and its binding poses in zinc-containing matrix metalloproteinase-9 (MMP-9). As demonstrated, the present QM/MM second-order generalized ensemble sampling technique enables feasible usage of the QM/MM model to sample molecular conformations in condensed environments.

A practical synthesis of differentially-protected cis-1,2-cyclopentanedithiols and cis-3,4-pyrrolidinedithiols

A practical method for the synthesis of cis-1,2-cyclopentanedithiols and cis-3,4-pyrrolidinedithiols with differentially protected sulfurs, needed for the design of new metal-chelating ligands, has been developed.

Matrix metalloproteinase inhibitors based on the 3-mercaptopyrrolidine core.

New series of pyrrolidine mercaptosulfide, 2-mercaptocyclopentane arylsulfonamide, and 3-mercapto-4-arylsulfonamidopyrrolidine matrix metalloproteinase inhibitors (MMPIs) were designed, synthesized, and evaluated. Exhibiting unique properties over other MMPIs (e.g., hydroxamates), these newly reported compounds are capable of modulating activities of several MMPs in the low nanomolar range, including MMP-2 (~2 to 50 nM), MMP-13 (~2 to 50 nM), and MMP-14 (~4 to 60 nM). Additionally these compounds are selective to intermediate- and deep-pocket MMPs but not shallow-pocketed MMPs (e.g., MMP-1, ~850 to >50,000 nM; MMP-7, ~4000 to >25,000 nM). Our previous work with the mercaptosulfide functionality attached to both cyclopentane and pyrrolidine frameworks demonstrated that the cis-(3S,4R)-stereochemistry was optimal for all of the MMPs tested. However, in our newest compounds an interesting shift of preference to the trans form of the mercaptosulfonamides was observed with increased oxidative stability and biological compatibility. We also report several kinetic and biological characteristics showing that these compounds may be used to probe the mechanistic activities of MMPs in disease.

A new synthetic matrix metalloproteinase inhibitor reduces human mesenchymal stem cell adipogenesis

Development of adipose tissue requires the differentiation of less specialized cells, such as human mesenchymal stem cells (hMSCs), into adipocytes. Since matrix metalloproteinases (MMPs) play critical roles in the cell differentiation process, we conducted investigations to determine if a novel mercaptosulfonamide-based MMP inhibitor (MMPI), YHJ-7-52, could affect hMSC adipogenic differentiation and lipid accumulation. Enzyme inhibition assays, adipogenic differentiation experiments, and quantitative PCR methods were employed to characterize this inhibitor and determine its effect upon adipogenesis. YHJ-7-52 reduced lipid accumulation in differentiated cells by comparable amounts as a potent hydroxamate MMPI, GM6001. However, YHJ-7-82, a non-inhibitory structural analog of YHJ-7-52, in which the zinc-binding thiol group is replaced by a hydroxyl group, had no effect on adipogenesis. The two MMPIs (YHJ-7-52 and GM6001) were also as effective in reducing lipid accumulation in differentiated cells as T0070907, an antagonist of peroxisome-proliferator activated receptor gamma (PPAR-gamma), at a similar concentration. PPAR-gamma is a typical adipogenic marker and a key regulatory protein for the transition of preadiopocyte to adipocyte. Moreover, MMP inhibition was able to suppress lipid accumulation in cells co-treated with Troglitazone, a PPAR-gamma agonist. Our results indicate that MMP inhibitors may be used as molecular tools for adipogenesis and obesity treatment research.