Zinc-binding groups modulate selective inhibition of MMPs

Abstract

The need for selective matrix metalloproteinase (MMP) inhibition is of interest because of the range of pathologies mediated by different MMP isoforms. The development of more selective MMP inhibitors (MMPi) may help to overcome some of the undesired side effects that have hindered the clinical success of these compounds. In an effort to devise new approaches to selective inhibitors, herein we describe several novel MMPi and show that their selectivity is dependent on the nature of the zinc-binding group (ZBG). This is in contrast to most current MMPi, which obtain isoform selectivity solely from the peptidomimetic backbone portion of the compound. In the present study, six different hydroxypyrone and hydroxypyridinone ZBGs were appended to a common biphenyl backbone and the inhibition efficiency of each inhibitor was determined in vitro (IC(50) values) against MMP-1, -2, -3, -7, -8, -9, -12, and -13. The results show that the selectivity profile of each inhibitor is different as a result of the various ZBGs. Computational modeling studies were used to explain some trends in the observed selectivity profiles. To assess the importance of the ZBG in a biological model, two of the semiselective, potent MMPi (and one control) were evaluated using an isolated perfused rat heart system. Hearts were subjected to ischemia reperfusion injury, and recovery of contractile function was examined. In this model, only one of the two MMPi showed significant and sustained heart recovery, demonstrating that the choice of ZBG can have a significant effect in a relevant pathophysiological endpoint.

Figure 1

Structures of the inhibitors examined in this study (1–7). An abbreviated name for each inhibitor is provided in parentheses. Compounds 6 and 7 are negative controls, as they do not contain a high affinity ZBG. The numbering scheme used for the pyrone ring system is shown on the left.

Figure 2

Dixon plot of MMP-12 with different substrate concentrations (10 µm=filled circles, 5 µm=open squares, 2.5 µm=open diamonds, 1.67 µm=crosses, 1 µm=plus signs) against varying concentrations of MMPi 1 (1/v in mmol⁻¹min mg, [I] in nm).

Figure 3

Recovery of rate pressure product (RPP) in isolated rat hearts subjected to global no-flow ischemia (GNFI) and 30 min reperfusion in the absence (controls) and presence of: 5 µm PY-2 (compound 2, upper left), 5 µm 1, 2-HOPO-2 (compound 1, upper right), and 5 µm PICO-2 (compound 6, lower left). A direct comparison of the recovery results from PY-2 and 1, 2-HOPO-2 are shown in the panel at the lower right. Note, at equal concentrations, 1, 2-HOPO-2 confers a higher recovery of contractile function than PY-2 during reperfusion. All MMPi were present in the perfusion buffer throughout the experiment. *P<0.05, **P<0.01.

Figure 4

Graph of IC₅₀ values (on y-axis) from Table 1 plotted on a logarithmic scale for inhibitors 1–5. Each MMP is color coded as shown in the legend (MMP-2 dark blue; MMP-3 turquoise; MMP-8 gray; MMP-12 green; MMP-13 mauve). The top plot shows all of the MMPs tested and the bottom plot shows only the intermediate and deep S1′ pocket MMPs.

Figure 5

Three different views comparing the binding of MMPi 3 (left) and 5 (right) in the MMP-3 active site. Zn²⁺ ions are shown as magenta spheres, the MMPi in ball-and-stick colored by atom, and the protein in gray. The three views highlight the steric clash of the N-methyl group on compound 5 with the protein. Val 163 (shown as sticks in the bottom views) is the residue that conflicts with the N-methyl substituent.