Correlation between the Affinity of Flavonoids Binding to the Cytosolic Site of Leishmania tropica Multidrug Transporter and Their Efficiency To Revert Parasite Resistance to Daunomycin

Abstract

The C-terminal nucleotide-binding domain (NBD2) of a P-glycoprotein-like transporter, encoded by the ltrmdr1 gene in Leishmania tropica and involved in parasite multidrug resistance (MDR), was overexpressed in Escherichia coli as a hexahistidine tagged protein and purified. The L. tropica recombinant domain efficiently bound fluorescent derivatives of ATP, the hydrophobic steroid analogue RU 486, and different classes of flavonoids with the following efficiency: flavone > flavanone > isoflavone > glucorhamnosyl-flavone > chromone. The affinity for flavones was dependent on the presence of hydroxyl groups at positions 5 and 3 and was further increased by a hydrophobic 1,1-dimethylallyl substituent at position 8. When flow cytometry was used to measure daunomycin accumulation in a MDR L. tropica line, a reversing effect was observed with flavones such as dimethylallyl-kaempferide at low concentration or apigenin at higher concentration, but neither with the glucorhamnosyl derivative rutin nor with the isoflavone genistein. The in vivo reversing effect of dimethylallyl-kaempferide was correlated to a high inhibition of MDR cell growth in the presence of daunomycin. The results suggest that flavone inhibition of both daunomycin efflux and parasite growth in the presence of the drug correlates to direct binding of the compound to cytosolic domain of the P-glycoprotein-like transporter.

A Quantitative Structure-Activity Relationship for the Modulation Effects of Flavonoids on P-Glycoprotein-Mediated Transport

“…This conforms to results reported by Conseil et al and Boumendjel et al who used the dissociation constant (K d ) toward NBD2 as a dependent variable. [10][11][12][13] A plot of the predicted percent modulation effect from the respective QSAR equations (the first three equations in Table 2) against observed values, given in Fig. 4, shows that the predicted percent inhibition values for all flavonoids were highly correlated with the observed values and Ͻ2-fold off from the observed percent inhibition values observed for the Y2 level, suggesting that this model does not have a tendency to over-or under-predict the percent inhibition values and thus is useful for predicting percent inhibition values of flavonoids.…”

“…In fact, both nonprenylated flavonoids and prenylated derivatives were found to interact with the hydrophobic region or outside the NBDs. [10][11][12] Based on measurements of binding affinity to-ward mouse NBD2 of P-gp, the structure-activity relationship analysis of flavonoid-P-gp concluded that flavonols, chalcones, and flavones are the most active, and flavanones and isoflavones have lower activities, and the presence of the 5-OH group, 3-OH group, and two or three double bonds are required for activity. 10,13) A quantitative structure-activity relationship (QSAR) analysis by stepwise regression was performed to construct the equation log(1/K d )ϭ0.335c log PϪ 0.007H f Ϫ0.114N lc ϩ4.342, (r 2 ϭ0.742; K d , dissociation constant toward NBD2; c log P, calculated logarithm of the octanol/water partition coefficient; H f , heat of formation; and N lc , number of hydrogen-suppressed atoms of the longest chain length) for quantitatively evaluating the most significant structural factors on the interaction between flavonoids and P-gp.…”

HPLC-DAD phenolic profile, cytotoxic and anti-kinetoplastidae activity ofMelissa officinalis

“…Other biochemical mechanisms include the inhibition of the p-glycoproteinlike transporter (Perez-Victoria et al 1999) or modulation of protein phosphorylation on the SPK89 protein kinase in trypanosomes (Gale et al 1994). Corroborating with what was previously mentioned; a more recent study showed that quercetin has a pro-oxidant effect, as it acted to produce reactive oxygen species, against the amastigote forms of Leishmania amazonensis, leading to dysfunction of its mitochondrial membrane (FonsecaSilva et al 2013).…”

Antileishmanial activity and evaluation of the mechanism of action of strychnobiflavone flavonoid isolated from Strychnos pseudoquina against Leishmania infantum

“…They are a promising new class of immune modulators for Leishmania spp. and have proven to bind to the nucleotide-binding site of MDR proteins with a concomitant increase in intracellular drug accumulation (Pérez-Victoria et al, 1999). Additionally, flavones potentiate the antibiotics of berbine and norfloxacin in Staphylococcus aureus, as well as artemisinin in Plasmodium falciparum (Liu et al, 1992), indicating their potential combined use within a chemotherapeutic regimen (Mead & McNair, 2006).…”