In Silico Identification and In Vitro Evaluation of Natural Inhibitors of Leishmania major Pteridine Reductase I

Abstract

In a continuation of our computational efforts to find new natural inhibitors of a variety of target enzymes from parasites causing neglected tropical diseases (NTDs), we now report on 15 natural products (NPs) that we have identified as inhibitors of Leishmania major pteridine reductase I (LmPTR1) through a combination of in silico and in vitro investigations. Pteridine reductase (PTR1) is an enzyme of the trypanosomatid parasites’ peculiar folate metabolism, and has previously been validated as a drug target. Initially, pharmacophore queries were created based on four 3D structures of LmPTR1 using co-crystallized known inhibitors as templates. Each of the pharmacophore queries was used to virtually screen a database of 1100 commercially available natural products. The resulting hits were submitted to molecular docking analyses in the substrate binding site of the respective protein structures used for the pharmacophore design. This approach led to the in silico identification of a total of 18 NPs with predicted binding affinity to LmPTR1. These compounds were subsequently tested in vitro for inhibitory activity towards recombinant LmPTR1 in a spectrophotometric inhibition assay. Fifteen out of the 18 tested compounds (hit rate = 83%) showed significant inhibitory activity against LmPTR1 when tested at a concentration of 50 µM. The IC50 values were determined for the six NPs that inhibited the target enzyme by more than 50% at 50 µM, with sophoraflavanone G being the most active compound tested (IC50 = 19.2 µM). The NPs identified and evaluated in the present study may represent promising lead structures for the further rational drug design of more potent inhibitors against LmPTR1.

Sesquiterpene Lactones with Dual Inhibitory Activity against the Trypanosoma brucei Pteridine Reductase 1 and Dihydrofolate Reductase

“…Both Tb DHFR and Tb PTR1 were obtained by recombinant expression in E. coli and enzyme inhibition assays established based on previous reports [ 23 , 24 , 25 ].…”

In-Vitro Activity of Silybin and Related Flavonolignans against Leishmania infantum and L. donovani

“…Mechanism of action of flavonolignans is not precisely known. Besides their antioxidant role, other potential targets have been indicated including the inhibition and modulation of drug transporters [ 15 , 29 , 30 ], suppression of cellular inflammation [ 17 ], stimulation of protein synthesis [ 16 ], and significant inhibitory activity in vitro against L. major pteridine reductase I (LmPTR1) [ 31 ]. In our case, we found that oxidized stereoisomers displayed a higher toxicity for promastigotes, Mφ and intracellular amastigotes than the parent molecule.…”

In-Silico Identification of the Best Compound Against <i>Leishmania infantum</i>: High Throughput Screening of All FDA Approved Drugs

“…The "in-silico" analysis which stands for computer-based biological experiments, is a state-of-the-art and accurate method to discover exclusive bioactive compounds, which may represent novel metabolic pathways and/or a powerful affinity to a certain target (42). In this sense, several studies have identified unprecedented molecules for various drug targets in Leishmania, such as pteridine reductase1, tryparedoxin peroxidase and sterol biosynthesis (43)(44)(45)(46)(47)(48), as well as in other single-celled eukaryotes enclosing Toxoplasma gondii, Cryptosporidium hominis, Plasmodium and Trypanosoma cruzi (49)(50)(51). Current in-silico investigation was aimed to screen and predict the anti-leishmanial potency of 3358 FDA-approved compounds against both drug targets in L. infantum in comparison to amphotericin B and glucantime for the discovery of new biochemical molecules.…”