In Vitro Screening to Identify Anti- Toxoplasma Compounds and In Silico Modeling for Bioactivities and Toxicity

Abstract

Toxoplasmosis, which affects more than a billion people worldwide, is a common parasitic infection caused by the obligate intracellular parasite, Toxoplasmagondii. Current treatment strategies have several limitations, including unwanted side effects and poor efficacy. Therefore, newer therapies are needed for toxoplasmosis. Drug repurposing and screening of a vast array of natural and/or synthetic compounds is a viable option for antiparasitic drug discovery. In this study, we screened 62 compounds comprising natural products (NPs) and FDA-approved (FDA) drugs, to identify the hit compounds that suppress the growth of T. gondii. To determine the parasite inhibitory potential of the compounds, host mammalian cells were infected with a transgenic T. gondii strain, and the viability of the parasite was evaluated by luminescence. Of the 62 compounds, tubericidin, sulfuretin, peruvoside, resveratrol, narasin and diacetoxyscirpenol of the natural product isolates, as well as bortezonib, 10-Hydroxycamtothecin, mebendazole, niflumic acid, clindamycin HCl, mecamylamine, chloroquine, mitomycin C, fenbendazole, daunorubicin, atropine, and cerivastatin of FDA molecules were identified as "hits" with ≥ 40 percent anti-parasite action. Additionally, mitomycin C, radicicol, naringenin, gitoxigenin, menadione, botulin, genistin, homobutein, and gelsemin HCl of the natural product isolates, as well as lomofungin, cyclocytidine, prazosin HCl, cerivastatin, camptothecin, flufenamic acid, atropine, daunorubicin, and fenbendazole of the FDA compounds exhibited cytotoxic activity, reducing the host viability by ≥ 30 percent. Our findings not only support the prospects of drug repurposing, but also indicate that screening a vast array of molecules may provide viable sources of alternative therapies for parasitic infection.

Keywords: Antiparasite; Drug discovery; Library of chemical compounds; Medicinal biochemistry; Parasitic infection.

Figure 1

Growth inhibition of Toxoplasma gondii RH-2F strain by natural product isolates at a concentration of 1 μg/mL. DMSO (0.5%) control was calculated as 0% inhibition, and 200 μg/mL pyrimethamine positive drug control was calculated as 100% growth inhibition. Data are presented as mean values of three replicates ± standard errors of the mean.

Figure 2

Growth inhibition of Toxoplasma gondii RH-2F strain by FDA-approved compounds at a concentration of 1.5 μg/mL. DMSO (0.5%) control was calculated as 0% inhibition, and 200 μg/mL pyrimethamine positive drug control was calculated as 100% growth inhibition. Data are presented as mean values of three replicates ± standard errors of the mean.

Figure 3

Viability of host cell (human foreskin fibroblast) by natural product isolates at 2 μg/mL. DMSO (0.5%) control was calculated as 100% cell viability. Data are presented as mean values of three replicates ± standard errors of the mean.

Figure 4

Viability of host cell (human foreskin fibroblast) by FDA-approved compounds at 3 μg/mL. DMSO (0.5%) control was calculated as 100% cell viability. Data are presented as mean values of three replicates ± standard errors of the mean.

Figure 5

Chemical structures of hit compounds from the natural product isolates.

Figure 6

Chemical structures of hit compounds from the FDA-approved drugs.

Figure 7

Distribution of active natural product compounds based on (A) bioactivity, (B) functional groups, and (C) treatment indications for FDA-approved drugs.