MDN-0185, an Antiplasmodial Polycyclic Xanthone Isolated from <i>Micromonospora</i> sp. CA-256353

A potent antiplasmodial polycyclic xanthone, MDN-0185 (<b>1</b>), was isolated from an unidentified species of the genus <i>Micromonospora</i>. The planar structure of <b>1</b> was established as a seven-ring polycyclic xanthone with partial structures very similar to two known natural products, namely, xantholipin and Sch 54445. Using ROESY correlations, the relative stereochemistry of the two independent stereoclusters of compound <b>1</b> could be determined. Mosher analysis and comparison of the specific rotation of compound <b>1</b> with that of xantholipin allowed the determination of its absolute configuration. Compound <b>1</b> exhibited an IC₅₀ of 9 nM against <i>Plasmodium falciparum</i> 3D7 parasites

Quantification of intracellular ATP levels.

(A) ATP standard curve used for interpolating RLU read outs. (B) T. cruzi epimastigotes were treated with strasseriolide C for 72 hours at increasing concentrations, and ATP content was determined using Cell-Titer Glo luminescent viability assay and normalized by the number of cells per well. Results correspond to the mean of three replicates for each condition. (TIF)</p

Rate of action against <i>T</i>. <i>cruzi</i> trypomastigotes.

(A) Effect of different concentrations of strasseriolide C, benznidazole and nifurtimox on T. cruzi trypomastigotes viability. Loss of viability upon 24, 48 and 72 hours drug treatment was measured by the luminescent method at 1/2 serial concentrations ranging from 50 μM strasseriolide and benznidazole and from 10 μM for nifurtimox. Results correspond to the mean value of 3 replicates and error bars represent the standard deviation. (B) Maximum percentage activity against extracellular forms of T. cruzi reached following 24 and 48 hours treatment with 50 μM strasseriolide C (blue bars), 50 μM benznidazole (green bars), 10 μM nifurtimox (red bars) or 2 μM posaconazole (purple bars) in rate of kill experiments with trypomastigote forms of the parasite.</p

Cell cycle progression of <i>T</i>. <i>cruzi</i> epimastigotes after 72-hour drug exposure.

(A) Fluorescence microscopy with DAPI staining. Bars represent the mean (± SD) calculated from two independent experiments accounting for at least 80 cells per condition respectively. (B) FACS analysis with propidium iodide staining. Bars represent the mean (± SD) calculated from two independent experiments and analyzed with Flow Jo software. (C) Overlay of fluorescence histograms obtained for control and treated fixed parasites stained with propidium iodide.</p

Rate of action against <i>T</i>. <i>cruzi</i> amastigotes.

(A) Time-dependent correlation between compound dose and T. cruzi amastigote growth inhibition upon 24, 48 and 72 hours exposure plotted as 10-point dose-response curves. Dose-response data obtained for posaconazole after 24, 48 and 72 h exposure at higher concentrations is indicated. (B) Maximum percentage activity against intracellular forms of T. cruzi obtained after 24, 48 and 72 hours treatment with 50 μM strasseriolide C (blue bars), 50 μM benznidazole (green bars), 20 μM nifurtimox (red bars) or 100 nM posaconazole (purple bars) in rate of kill experiments with amastigote forms of the parasite.</p

Fig 2 -

Dose-response curves for T. cruzi intracellular amastigotes after 96-hour treatment with strasseriolides A-D. Growth inhibition has been quantified by CPRG colorimetric read out following the β-D-galactosidase transgenic T. cruzi assay. Bars represent the mean (± SD) at each of the 16 concentration points calculated from a minimum of three biological replicates.</p

S1 Fig -

HPLC-UV (210 nm) traces of pure strasseriolides A-D. (TIF)</p

Excel spreadsheet containing, in separate sheets, all numerical values for Figs 1, 2A, 2B, 3A, 3B, 4A, 4B, 5A, 6, S2B, S3A, S3B, S4A and S4B.

Excel spreadsheet containing, in separate sheets, all numerical values for Figs 1, 2A, 2B, 3A, 3B, 4A, 4B, 5A, 6, S2B, S3A, S3B, S4A and S4B.</p

Time-dependent resazurin reduction by <i>T</i>. <i>cruzi</i> trypomastigotes at different cell densities.

Trypomastigotes were obtained as indicated in the methods section and prior to the resazurin assay, were incubated for 24 (panel A) and 48 hours (panel B) at 37°C. Fluorescence was measured after 2 hours (circles) and 24 hours (triangles) of incubation with resazurin. (TIF)</p