Design, synthesis and antimalarial activity of novel, quinoline-Based, zinc metallo-aminopeptidase inhibitors
Abstract
A new series of antimalarials consisting in 45 potential inhibitors of PfA-M1 is described.
Introduction
Malaria, caused by the parasite Plasmodium falciparum, is still prevalent and lethal in many countries.1 Chloroquine (CQ), exerting its antimalarial activity by inhibiting haemozoin formation in the food vacuole of the parasite, has been the standard drug for many decades. Unfortunately, the spread of resistant P. falciparum to this molecule has created an urgent need to develop new antimalarial treatments.2 In a project to find new quinoline-based antimalarials that would not induce resistance, unlike CQ, we have designed several compounds displaying a piperazine linker. Several of these compounds were active on the CQ-resistant strain FcB1.3
The recent publication of the genome of the parasite opens the opportunities of a better understanding of the biology of the parasite.4 This will hopefully result in the discovery of new specific targets to tackle with drugs. Proteases expressed in the erythrocytic stage of P. falciparum, for example, can be considered as interesting targets for the design of antimalarials.5, 6
In particular, PfA-M1,7 a neutral zinc-aminopeptidase (M1 family), inhibited by the classical aminopeptidase inhibitor bestatin, has been proposed:
(1) to hydrolyze the small peptides generated in the food vacuole, during hemoglobin digestion by acidic endopeptidases (aspartyl, cysteine and metallo-endopeptidases), into amino-acids at the level of the cytoplasm, and: (2) to play a role in the erythrocyte re-invasion by the parasite.8 PfA-M1 shares a maximal homology in the active site region (44%) with two human proteases: the human aminopeptidase-N and leukotriene A4 hydrolase.9
It is the only aminopeptidase of P. falciparum that has been purified and biochemically characterized. This enables the design and testing of potential inhibitors. Such compounds would help understanding the role of this enzyme and validate it as a therapeutic target.
In a project aiming at decreasing the potential for resistance occurrence of our quinoline-based antimalarials, we have designed dual inhibitors that would inhibit both the haemozoin formation and one of the proteases potentially involved in globin digestion. Such a strategy has been developed by Avery for the design of malarial cysteine proteases inhibitors based on mefloquine and chloroquine.10 The screening of several of our in-house quinoline-based compounds allowed us to identify compound 1 as an interesting inhibitor of PfA-M1, that served as a starting point for analoguing (Fig. 1).
We report here the design and parallel synthesis of a library of 45 non-peptidic analogues of 1, and its biological evaluation on PfA-M1. Preliminary results on the antimalarial activity on FcB1 strain of the most active enzyme inhibitors, and preliminary specificity data on mammalian aminopeptidase N, are presented and discussed.
Section snippets
Chemistry
PfA-M1 is a zinc-metalloprotease, of the M1 family, preferably cleaving basic, hydrophobic, as well as aromatic amino-acids.8 Zn-chelating groups are critical for the inhibition of such enzymes.11 To fulfill this function, we incorporated at least a carboxylic acid or an hydroxamate group in our molecules. In order to mimic the side chain of a Leucine residue, an isobutyl group (Ibu), analogue of the methyl-cyclopropyl moiety in 1, was also added in our potential inhibitors.
The 45 analogues (5
Biological Assays
The analogues and the deprotected precursors were screened for their ability to inhibit PfA-M1 at 10 μM.14 Crude products displaying an inhibition percentage above 50% were selected for re-synthesis and IC50 determination on fully purified and controlled samples. The selected compounds were then tested for their ability to inhibit FcB1 parasite growth.15 The specificity against other aminopeptidases was evaluated in a model of mammalian aminopeptidase-N from porcine kidney.16
Conclusion
We have successfully designed non-peptidic inhibitors of PfA-M1, an aminopeptidase of P. falciparum. The quinoline moiety allows these compounds also to be inhibitors of haem detoxification, making these compounds potentially dual inhibitors of hemoglobin digestion. Yet the relative part of each mode of action needs to be determined. The three selected inhibitors 33–35 are active against CQ-resistant strain, thus validating their ability to cross membranes. The activity on PfA-M1, and the
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Cited by (56)
Antimalarial agents acting on hemoglobin degradation
2020, Antimalarial Agents: Design and Mechanism of ActionTwo cap residues in the S1 subsite of a Plasmodium falciparum M1-family aminopeptidase promote broad specificity and enhance catalysis
2017, Molecular and Biochemical ParasitologyTargeting the active sites of malarial proteases for antimalarial drug discovery: approaches, progress and challenges
2017, International Journal of Antimicrobial AgentsCitation Excerpt :They found a hydroxamate group to be more efficient than a carboxylate and therefore proposed the chelation of Zn2+ by the hydroxamate group as the main anchoring point. In addition, these researchers reported quinolone-class PfM1-AAP inhibitors [150]. The lead (compound 64; Fig. 22) exhibited IC50 values of 0.854 µM and 0.317 µM against PfM1-AAP and parasite growth, respectively.
KBE009: An antimalarial bestatin-like inhibitor of the Plasmodium falciparum M1 aminopeptidase discovered in an Ugi multicomponent reaction-derived peptidomimetic library
2017, Bioorganic and Medicinal ChemistryCitation Excerpt :The lack of correlation between the PfA-M1 inhibition in vitro and the antimalarial activity of the peptidomimetics listed in Table 3 is similar to the results reported by other authors for bestatin. The inhibitory potency of bestatin toward PfA-M1 is characterized by Ki values in the range 120–3800 nM.5,10,11,20,24 However, this compound inhibits the growth of P. falciparum in cultures with IC50 values from units to tenths of μM.2,5,10,20,25,27,28
Potent dual inhibitors of Plasmodium falciparum M1 and M17 aminopeptidases through optimization of S1 pocket interactions
2016, European Journal of Medicinal ChemistryHigh-level expression in Escherichia coli, purification and kinetic characterization of Plasmodium falciparum M1-aminopeptidase
2014, Protein Expression and PurificationCitation Excerpt :This enzyme, named PfAM1, has biological functions that are crucial for P. falciparum intra-erythrocytic stages inside the human host. PfAM1 is involved in the degradation of erythrocytic hemoglobin [1,5–11], trophozoite maturation [7,12,13], and reinvasion [7,9,11,13–15] and appears to be essential for the parasite during the erythrocytic stage [1,8,10,16–20]. Furthermore, inhibitors of PfAM1 block the in vitro growth of the parasite at micromolar concentrations [7,10,16,18–22], and one of them reduces infection in a murine malaria model [19].