Design, Synthesis And Activity Evaluation Of New Irreversible Myeloperoxidase Inhibitors Derived From Benzodioxole

The role of Myeloperoxidase (MPO) in the oxidative damages and the inflammatory syndromes is well documented. Thus, the inhibition of MPO in the circulation can be useful in the treatment of several inflammatory diseases. Some potent reversible MPO inhibitors derived from fluorotryptamine were published. In addition we have reported that the SSRI agent (paroxetine) can irreversibly inhibit MPO at low nanomolar range. With the docking experiments, the important chemical groups in both paroxetine and fluorotryptamine derivatives were determined and general structure of the new series was designed. This general structure consists of dioxole, aromatic ring Ar, hydrogen bond donor HBD and a space between HBD and Ar. Several modifications were applied to study the SAR of this series. These compounds were synthesized and tested in vitro. It is found that the IC50 of the compounds with amine are the lowest values among all the functional groups (IC50= 10-60 nM), that 5 carbons on the side chain give the best activity. Dioxole group is very important for the activity and the irreversibility. The in vitro test of these compounds on SERT improved the selectivity vs SERT.


Introduction
4 MPO, EC 1.11.2.2  The heme enzyme myeloperoxidase is a lysosomal protein that plays an important role in innate immunity system. It is expressed in neutrophils and stored in their azurophilic granules.  After phagocytosis of pathogens by the neutrophils, MPO produces a powerful oxidizing agent HOCl from H 2 O 2 and Clwhich leads to the oxidation (degradation) of biomolecules of pathogens in the phagosome.

97% yield
This compound was obtained by the the same procedure as for the amino compounds.
After the reaction was finished the reagent was evaporated.
Chain length: 5 carbons on the side chain gives the best activity for the compounds with amine group while for the amide and nitrile the best compounds are those with 4 carbons. IC 50 = 13 nM IC 50 = 52 nM IC 50 = 31 nM Functional group: the effect of the functional group is as following: -NH 2 > =NH> =N-> -CONH 2 > -CN> -OH> -Cl> -CH 3 . And =N + = has no activity.
Cyclic functional group: among piperazine, morpholine and pyrrolidine, the piperazine gives the best activity with the same activity of the compound with =NH.
Bridge: the best activity was shown when the bridge is ether. When the bridge is ester or amide the activity is lost. Docking could explain all the results except losing the activity in the compound without dioxole (dihydroxyl), the compound with ester bridge and the compound with amide bridge. The compounds that feature hydrogen bond or salt bridge with Glu102 have high potency DG= -22.9 kcal DG= -20.6 kcal DG= -13.1kcal In vitro test and SAR study SERT inhibition: in vitro test of all the synthetic compounds showed that these compounds have no activity on SERT, so our new inhibitors are selective for MPO. In order to improve the irreversible inhibitory effect of our new compounds on MPO, several concentrations of (the best new inhibitor, paroxetine and potent reversible inhibitor) were incubated with fixed amounts of MPO for 1h. After 1h, the activity of MPO was measured. It is found that when: the concentration of our new inhibitor is 50 times higher than this of MPO, the inhibition is 100%, the concentration of our paroxetine is 100 times higher than this of MPO, the inhibition is 100%. But with the reversible inhibitor, the inhibitory effect cannot reach at 100%.
 We developed the first potent irreversible inhibitors of MPO that inhibit the enzyme at nanomolar range. These inhibitors are derived from benzodioxole.
 The compounds that have amine on the side chain have the best activity.
 Five carbons between the bridge and the amine give the best activity.
 Ether group as a bridge between aromatic group and alkyl chain gives the best activity.
 The compound which has not dioxole reacts with both Compound I and Compound II of MPO in very fast way, so this molecule cannot cause accumulation of the inactive form of MPO (Compound II). This makes the compound with no activity.
 The most potent inhibitor among the synthesized compound has IC 50 of 13 nM.